Letters to the Editor
and previous results. For understanding the
chronological changes in TLR1 and 4-IR, we
would like to provide a summary of TLR2
and 4- IR in the dental pulp at different stages
(Table 1). The reason that it is difficult to pro-
vide statistical analysis is that it is impossible
to perform the same quantitative analyses in
tissue sections (present study) versus other
methods such as Western blots.
Regarding the request for additional in-
formation on qRT-PCR, we are happy to pro-
vide the primer information in Table 2; the
experimental methods are the same as previ-
ously reported by our group (2). We do agree
that future research could evaluate differential
expression in specific cell subsets.
In the present study, we evaluated pulpal
responses to dentin exposed to the oral
environment, with a focus on changes in the
expression of TLR 2 and TLR 4. We appreciate
the suggestion that future studies may benefit
from analyzing the relationship between the
functional T/B cells and the expressions of
TLR 2 and 4. However, the present results
do support the clinical implications that early
treatment of a carious lesion is a biologically
rational concept because it may augment the
innate immune responses to carious antigenic
challenges.
Nobuyuki Tani-Ishii
Department of Oral Medicine
Division of Endodontics
Kanagawa Dental College
Yokosuka, Japan
References
1. Mutoh, et al. J Endod 2009;35:975–80.
2. Mutoh, et al. J Endod 2007;33:1183–6.
Antibacterial Efficacy of MTAD Final
Rinse and Two Percent
Chlorhexidine Gel Medication in
Teeth with Apical Periodontitis
To the Editor:
I am writing this letter in response to
a clinical article that was published in
the November 2009 issue of Journal of End-
odontics (Volume 35, page 1483) to evaluate
the antimicrobial efficacy of BioPure MTAD
(Dentsply International, Tulsa, OK). Reading
this article raised a number of concerns that
highlight the challenges facing clinical studies
from a microbiological standpoint. In the in-
terest of brevity, I would like to draw attention
to Figure 2. As expected, 100% of the 1A (pre-
596 Letters to the Editor
treatment) samples showed bacterial growth.
The 1B samplings (post biomechanical in-
strumentation) showed approximately 30%
to 40% of teeth with remaining bacteria.
This finding is consistent with previous find-
ings in the literature (1–5). The microbial
findings in the 1C samples, however, raise
concerns about the likelihood of contamina-
tion. Nearly 40% more teeth showed bacterial
growth after receiving a final rinse of saline,
and there were approximately twice as many
teeth showing bacterial growth after receiving
a final rinse of BioPure MTAD. Although the
authors go through great lengths to discuss
the methodologies used in this study, they
fail to explain why the number of samples
that yielded growth doubled after the use of
BioPure MTAD. In fact, they state in the Dis-
cussion section that no significant difference
was observed in the proportion of positive-
growth samples. Regardless of what the
proven efficacy of BioPure MTAD might be
in eliminating bacteria from the root canal
system, at the very least, we would not expect
to find a 100% increase in the number of sam-
ples showing growth after its use. The reader
would have appreciated a more in depth dis-
cussion of these unexpected findings and the
validity of the findings. Perhaps a brief refer-
ence to the contaminated access samplings
(1A-C and 2A-C) as a source of root canal
contamination would have been appropriate.
In closing, perhaps the rationale to de-
velop a root canal irrigant that would compete
with sodium hypochlorite is a futile effort in
the first place. After all, bleach has proven rel-
atively effective, readily available, and cheap.
Nevertheless, although it did not eliminate
the use of sodium hypochlorite, the aim of
BioPure MTAD’s development and use was
to enhance debridement efforts during
endodontic procedures. Currently, it is the
only root canal irrigant that simultaneously
removes smear layer and bacteria from the
root canal system (6, 7). Before its Food
and Drug Administration approval and distri-
bution in the market, systematic studies were
conducted to evaluate its ability to remove
smear layer with minimal structural damage
to dentin and its antimicrobial efficacy (6,
8). Although studies should be conducted to
confirm or refute the initial studies, claims
that the initial findings cannot be repeated
can only be reserved for studies that duplicate
those methodologies.
Shahrokh Shabahang, DDS, MS, PhD
Loma Linda School of Dentistry
Loma Linda, CA, USA
References
1. Shabahang S, Torabinejad M. J Endod 2003;29:576–9.
2. Siqueira J, Rocas I, Favieri A, et al. J Endod
2000;26:331–4.
3. Sjogren U, Figdor D, Persson S, et al. Int Endod J
1997;30:297–306.
4. Sundqvist G, Figdor D, Persson S, et al. Oral Surg
Oral Med Oral Pathol Oral Radiol Endod 1998;85:
86–93.
5. Shuping GB, Ørstavik D, Sigurdsson A, et al. J Endod
2000;26:751–5.
6. Torabinejad M, Khademi AA, Babagoli J, et al. J Endod
2003;29:170.
7. Shabahang S, Pouresmail M, Torabinejad M. J Endod
2003;29:450–2.
8. Torabinajed M, Cho Y, Khademi AA, et al. J Endod
2003;29:233–9.
Reply to Dr Shabahang
To the Editor:
W e thank Dr Shabahang for his comments
related to the proportion of positive
cultures in root canal samples obtained be-
fore and after the application of MTAD in
our clinical trial (1). To briefly summarize
our in vivo results after the application of
MTAD, we reported a nonsignificant decrease
in bacterial counts using two vital microscopy
enumeration methods and a nonsignificant in-
crease in the proportion of bacterial growth in
culture. Dr Shabahang questions the validity of
the results, which he considers unexpected in
view of previous studies showing removal of
the smear layer and bacteria in vitro. He sug-
gests the possibility of contamination of the
samples and that the article did not adequately
discuss these results.
The previous in vitro studies have, in fact,
served as the rationale for our clinical study.
Because in vitro findings cannot be extrapo-
lated to the clinical situation, it is incumbent
on the endodontic research community to
conduct clinical trials of new treatment regi-
mens to assess their true efficacy. Our clinical
trial was conducted with methodologic rigor
intended to overcome some of the challenges
inherent to this area of research, including
reducing the risk of contamination. In our de-
tailed discussion of the methodology, we em-
phasized the limited reliability of culture
results (in fact, those were provided only as
a means to facilitate discussion of our results
in relation to previous clinical studies in which
culture has been the predominant methodol-
ogy). Considering their proven low accuracy,
speculations attempting to explain specific cul-
ture findings are futile. However, Dr Shaba-
hang’s speculation that the findings were
affected by contamination from the access
JOE — Volume 36, Number 4, April 2010
 Letters to the Editor

cavities can in all likelihood be refuted. First,
using the far more accurate outcome measure
of vital microscopy, there was a nonsignificant
decrease in bacterial counts after MTAD was
used. If contamination occurred, it should
have affected an increase in these counts as
well. Second, the actual step of applying the fi-
nal rinse with MTAD is the shortest and prob-
ably least prone to contamination. If indeed
our results were affected by contamination
from the access cavities, we would have ex-
pected this to impact on the findings at samples
1B (postinstrumentation) and to still see an
improvement after application of MTAD.
In summary, notwithstanding the in vitro
efficacy of MTAD, we stated that our findings
did not support its antibacterial efficacy
when applied clinically as in our study. Be-
cause no significant reduction in bacterial
counts (and positive cultures) was observed
after application of MTAD, we believe our
statement to be appropriate.
Gevik Malkhassian, DDS, MSc, FRCD(C),
on behalf of the co-authors
University of Toronto
Toronto, Ontario, Canada
References
1. Malkhassian, et al. J Endod 2009;35:1483–90.

JOE — Volume 36, Number 4, April 2010 Letters to the Editor 597