The hPDLCs are human fibroblasts with multifunctional tissue that provide physical, formative and remodeling, nutritional, and sensory functions [
22]. In the present study, the hPDLCs were treated with different concentrations of LPS from
P. gingivalis or
E. coli. Our data showed that LPS from both
P. gingivalis and
E. coli induced pro-inflammatory cytokine (IL-1β, TNF-α, IL-6, IL-8) mRNA expressions in a dose-dependent manner. However,
P. gingivalis-treated cells responded in a dose-dependent manner at lower doses when compared to
E. coli-treated cells (Fig.
2). The different levels of induced inflammation from the two sources may be attributed to molecular differences in the lipid components. The lipid A from
P. gingivalis LPS is a monophosphate type lacking a phosphate group in the 4′ position that does not contain a tetradecanoic acid, but rather long-chain fatty acids made up of only acyloxyl groups [
23]. It is believed that LPS from
P. gingivalis is recognized mainly by TLR2, while LPS from
E. coli is recognized mainly by TLR4 [
23‐
26]. Uehara et al. also reported that human gingival fibroblasts showed more robust mRNA expression of pro-inflammatory cytokines following TLR2 ligation (
Mycoplasma-type diacyl lipopeptide) compared to TLR4 ligation (
E. coli-type lipid A) [
27]. These results indicate that different sources of LPS might induce inflammation through different mechanisms.
In recent decades, many studies have shown that LLLT reduces inflammatory reactions both in vitro and in vivo. Correa et al. induced periodontitis in mice with LPS and found that LLLT (GaAs laser, 904 nm) diminished inflammatory cell migration in a dose-dependent manner, with an energy dose of 3 J/cm
2 identified as the most effective dose [
28]. Pires et al. reported a model of collagenase-induced tendinitis and demonstrated that LLLT (780 nm), at an energy dose of 7.7 J/cm
2, suppressed the expression of IL-6 [
29]. Boschi et al. reported that LLLT (InGaAlP laser, 660 nm) significantly reduced the expression of IL-6 and TNF-α [
30]. According to previous studies conducted in our lab [
31], LLLT (GaAlAs laser, 660 nm) showed the most effective suppression of inflammation at the optimal dose of 8 J/cm
2, which was used in this study. In the current study, we noted that LPS-challenged hPDLCs showed similar results to the aforementioned studies, suggesting that LLLT significantly suppressed the mRNA expression of pro-inflammatory cytokines (IL-1β, TNF-α, IL-6, IL-8), leading us to conclude that LLLT has an anti-inflammatory effect in hPDLCs (Fig.
3). In addition, we noticed that the anti-inflammatory effect of LLLT was neutralized when the cAMP inhibitor SQ22536 was used (Fig.
3). We also observed that the level of cAMP from hPDLCs was elevated by both forskolin (cAMP promoter) and LLLT but decreased when SQ22536 (cAMP inhibitor) was added (Fig.
4), consistent with previous study that the level of cAMP increased by approximately 3- to 4-fold following LLLT treatment of human adipose-derived stem cells [
31]. This finding suggests that LLLT can act to stimulate the level of cAMP. Other studies have shown that the elevation of intracellular cAMP levels inhibited the transcriptional activity of NF-κB, which is a crucial transcription factor in the regulation of inflammation [
32,
33]. Possible mechanisms that regulate of NF-κB activity include the ability of cAMP to manage IκB degradation and IKK activity as well as to change the composition of NF-κB dimers and thereby block transcription [
34]. Indeed, we observed that LLLT significantly inhibited the transcriptional activity of NF-κB in our previous study of LPS-stimulated human adipose-derived stem cells [
31]. In the present study, we treated LPS-stimulated hPDLCs with LLLT and the adenylyl cyclase inhibitor SQ22536 and observed that the inhibitory effect of LLLT on NF-κB transcriptional activity was significantly reduced (Fig.
5). Aimbire et al. [
35] also reported results similar to ours, showing that a low-level laser (660 nm) at an energy dose of 7.5 J/cm
2 inhibited NF-κB transcriptional activity and further reduced apoptotic gene expression.
Exploring LLLT’s role in regulating the induction of pro-inflammatory cytokine in periodontal pathology is important as it may lead to novel therapeutic approaches for periodontitis. This study has demonstrated that LLLT inhibits inflammation, induced by LPS from E. coli and P. gingivalis, through the cAMP/NF-ĸB signaling pathway in hPDLCs. Future research into the detailed regulation of LLLT on cAMP may be of great value in improving periodontal therapy.