Microbiome of periodontitis and peri-implantitis before and after therapy: Long-read 16S rRNA gene amplicon sequencing.

AIMS: The microbial profiles of peri-implantitis and periodontitis (PT) are inconclusive. The controversies mainly arise from the differences in sampling sites, targeted gene fragment, and microbiome analysis techniques. The objective of this study was to explore the microbiomes of peri-implantitis (PI), control implants (CI), PT and control teeth (CT), and the microbial change of PI after nonsurgical treatment (PIAT). METHODS: Twenty-two patients diagnosed with both PT and peri-implantitis were recruited. Clinical periodontal parameters and radiographic bone levels were recorded. In each patient, the subgingival and submucosal plaque samples were collected from sites with PI, CI, PT, CT, and PIAT. Microbiome diversity was analyzed by high-throughput amplicon sequencing using full-length of 16S rRNA gene by next generation sequencing. RESULTS: The 16S rRNA gene sequencing analysis revealed 512 OTUs in oral microbiome and 377 OTUs reached strain levels. The PI and PT groups possessed their own unique core microbiome. Treponema denticola was predominant in PI with probing depth of 8-10 mm. Interestingly, Thermovirga lienii DSM 17291 and Dialister invisus DSM 15470 were found to associate with PI. Nonsurgical treatment for peri-implantitis did not significantly alter the microbiome, except Rothia aeria. CONCLUSION: Our study suggests Treponemas species may play a pivotal role in peri-implantitis. Nonsurgical treatment did not exert a major influence on the peri-implantitis microbiome in short-term follow-up. PT and peri-implantitis possess the unique microbiome profiles, and different therapeutic strategies may be suggested in the future.

Oral microbiome and serological analyses on association of Alzheimer's disease and periodontitis.

OBJECTIVE: To investigate the association between Alzheimer's disease (AD) and periodontitis in the aspects of periodontal status, serological markers, and oral microbiome. MATERIALS AND METHODS: Twenty AD and 20 healthy subjects were enrolled in this age- and gender-matched case-control study. Clinical periodontal parameters and serum biomarkers, including amyloid ß42 (Aß42 ), Tau, phosphorylated Tau (pTau), triglyceride, pro-inflammatory cytokines, and anti-Porphyromonas gingivalis lipopolysaccharide (LPS) antibody were examined. The saliva samples were analyzed for oral microbiome composition. RESULTS: Alzheimer's disease patients with Clinical Dementia Rating (CDR) ≥1 exhibited significantly more clinical attachment loss (CAL) than those with lower CDR. The levels of serum Tau protein, hsCRP and anti-P. gingivalis LPS antibody were markedly elevated in the AD group compared with the control group. Serum pTau protein level was positively correlated with anti-P. gingivalis LPS antibody titer. Moreover, the increased abundances of Capnocytophaga sp ora clone DZ074, Eubacterium infirmum, Prevotella buccae, and Selenomonas artemidis were detected in the AD group. Interestingly, serum levels of Aß42, pTau, and anti-P. gingivalis LPS antibody were strongly related to the gene upregulation in human pathogen septicemia. CONCLUSIONS: Our study suggested the association of periodontal infection and oral microbiome with AD. Further large-scale studies with longitudinal follow-up are warranted.

Association between periodontitis and spontaneous abortion: A case-control study.

BACKGROUND: Spontaneous abortion, or miscarriage, is a complication of pregnancy which can severely affect women both physically and psychologically. We investigated the associations of periodontitis and periodontopathic bacteria with spontaneous abortion. METHODS: We conducted a matched case-control study in two tertiary hospitals in Khon Kaen, Thailand. Cases were 85 women with spontaneous abortion at <20 weeks of gestation matched to 85 controls on age, gestational age, and hospital. Full-mouth periodontal examinations were performed. Periodontitis was defined as at least one site with probing depth ≥5 mm and clinical attachment level ≥2 mm at the same site. Subgingival plaque samples were collected to determine the levels of Porphyromonas gingivalis, Tannerella forsythia, and Fusobacterium nucleatum using real time polymerase chain reaction. RESULTS: The cases were significantly more likely to have periodontitis (50.6%) than the controls (21.2%; P = 0.007). Conditional logistic regression revealed a crude odds ratio (OR) of 4.1 for the association between periodontitis and spontaneous abortion (95% confidence interval [CI] = 1.9-8.9, P = 0.001). The OR decreased, but was still significant, after controlling for previous miscarriage (OR = 3.3, 95% CI = 1.4-7.8, P = 0.006). There was no significant association between the levels of periodontopathic bacteria and spontaneous abortion. Increased levels of P. gingivalis and F. nucleatum were associated with periodontitis in both case and control groups. Association between increased T. forsythia levels and periodontitis was observed only in the case women. CONCLUSIONS: Periodontitis was more common in women with spontaneous abortions as compared with matched controls. Levels of periodontopathic bacteria was not associated with spontaneous abortion in this population.

ß2-Glycoprotein I-Dependent Anti-Cardiolipin Antibodies Associated With Periodontitis in Patients With Systemic Lupus Erythematosus.

BACKGROUND: It was reported that patients with systemic lupus erythematosus (SLE) exhibited increased levels of anticardiolipin (anti-CL) antibodies, a class of antiphospholipid antibodies associated with thrombosis. ß2-glycoprotein I (ß2GPI) has been considered as the actual target antigen for anti-CL antibodies. This study investigates the association of periodontal infection with anti-CL antibodies in patients with SLE. METHODS: Fifty-three SLE female patients and 56 healthy female volunteers were recruited in this case-control study. All participants received periodontal examinations. The presence of Porphyromonas gingivalis and Treponema denticola in saliva and plaque samples was detected by polymerase chain reaction. Levels of serum anti-CL and anti-ß2GPI antibodies were examined using enzyme-linked immunosorbent assay. RESULTS: Patients with SLE exhibited more periodontal attachment loss and increased titers of serum anti-CL and anti-ß2GPI antibodies compared with healthy controls. Patients with active SLE who harbored P. gingivalis or P. gingivalis together with T. denticola intraorally exhibited significantly higher anti-CL and anti-ß2GPI antibodies than those without these bacteria. Anti-CL and anti-ß2GPI antibody levels correlated positively with clinical attachment level. Furthermore, increased anti-ß2GPI antibody levels were significantly associated with C-reactive protein and erythrocyte sedimentation rate. CONCLUSIONS: Elevated anti-CL and anti-ß2GPI antibody levels were associated with periodontopathic bacteria and periodontal breakdown in patients with SLE. Periodontitis might be a modifiable risk factor for SLE.

Induction of toll-like receptor expression by Porphyromonas gingivalis.

BACKGROUND: Toll-like receptors (TLRs) play pivotal roles in host immune responses and have been suggested to be involved in the development of many infectious diseases. In this study, the mRNA expression levels of TLR2, TLR4, and TLR9 and their relationship with periodontopathic bacteria in periodontal tissue are examined. Furthermore, the mechanism of TLR induction by Porphyromonas gingivalis is investigated in human gingival fibroblasts (HGFs). METHODS: Gingival tissue and subgingival plaque samples were collected from 19 patients with chronic periodontitis (CP) and 16 control individuals without periodontitis. Gene expression levels in the tissues and in HGFs were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). The numbers of periodontopathic bacteria were determined by quantitative real-time PCR. RESULTS: The expression levels of TLR2 and TLR9 were significantly higher in the tissues of patients with CP compared to the tissues of control individuals. The mRNA levels of TLR2 and TLR9, but not TLR4, were positively correlated with the number of P. gingivalis in subgingival plaque. P. gingivalis sonicated extract, P. gingivalis lipopolysaccharide, P. gingivalis DNA, and tumor necrosis factor-α(TNF-α) could significantly upregulate the mRNA expression of TLR2 in HGFs. Furthermore, P. gingivalis-mediated TLR2 expression was suppressed by TNF-α antibody. CONCLUSIONS: This study suggests that P. gingivalis infection induces TLR2 and TLR9 upregulation in patients with CP. P. gingivalis-induced TLR2 expression in HGFs is partially dependent on TNF-α and may lead to sensitization of HGFs to bacterial components encountered in the periodontal microenvironment.

Periodontitis and gestational diabetes mellitus in non-smoking females.

BACKGROUND: Chronic inflammation has been implicated in the pathogenesis of gestational diabetes mellitus (GDM). Periodontal disease is associated with increased levels of inflammatory mediators and may be a risk factor for GDM. The authors aimed to examine the association between periodontitis and GDM among non-smoking pregnant females. METHODS: This case-control study included 50 females who were diagnosed with GDM and 50 age- and hospital-matched females without diabetes in Khon Kaen, Thailand. Full-mouth periodontal examinations were performed during pregnancy by two calibrated dentists who were unaware of the case-control status. Periodontitis was defined as ≥1 site with probing depth (PD) ≥5 mm and clinical attachment level (CAL) ≥2 mm at the same site. Serum samples were collected to measure C-reactive protein (CRP), tumor necrosis factor-α, and interleukin-6 levels. Analyses were performed using conditional logistic regression. RESULTS: Fifty percent of the case females had periodontitis compared to 26% of the controls. Females with GDM had significantly higher mean PD and CAL, more sites with bleeding on probing, and increased levels of CRP compared to the controls. Periodontitis was significantly associated with GDM (odds ratio = 3.00, 95% confidence interval = 1.19 to 7.56). The association remained significant with additional adjustment for family history of diabetes, prepregnancy body mass index, and weight gain during pregnancy. CONCLUSIONS: The results suggest that periodontitis is associated with GDM. Therefore, clinicians should assess periodontal conditions of pregnant females.

Modulation of Wnt5a expression by periodontopathic bacteria.

Wingless proteins, termed Wnt, are involved in embryonic development, blood cell differentiation, and tumorigenesis. In mammalian hematopoiesis, Wnt signaling is essential for stem-cell homeostasis and lymphocyte differentiation. Recent studies have suggested that these molecules are associated with cardiovascular diseases, rheumatoid arthritis, and osteoarthritis. Furthermore, Wnt5a signaling is essential for the general inflammatory response of human macrophages. Periodontitis is a chronic inflammatory disease caused by gram-negative periodontopathic bacteria and the resultant host immune response. Periodontitis is characterized by loss of tooth-supporting structures and alveolar bone resorption. There have been no previous reports on Wnt5a expression in periodontitis tissue, and only few study reported the molecular mechanisms of Wnt5a expression in LPS-stimulated monocytic cells. Using RT-PCR, we demonstrated that Wnt5a mRNA expression was up-regulated in chronic periodontitis tissue as compared to healthy control tissue. P. gingivalis LPS induced Wnt5a mRNA in the human monocytic cell line THP-1 with a peak at 4 hrs after stimulation. P. gingivalis LPS induced higher up-regulation of Wnt5a mRNA than E. coli LPS. The LPS receptors TLR2 and TLR4 were equally expressed on the surface of THP-1 cells. P. gingivalis LPS induced IκBα degradation and was able to increase the NF-κB binding activity to DNA. P. gingivalis LPS-induced Wnt5a expression was inhibited by NF-κB inhibitors, suggesting NF-κB involvement. Furthermore, IFN-γ synergistically enhanced the P. gingivalis LPS-induced production of Wnt5a. Pharmacological investigation and siRNA experiments showed that STAT1 was important for P. gingivalis LPS-induced Wnt5a expression. These results suggest that the modulation of Wnt5a expression by P. gingivalis may play an important role in the periodontal inflammatory process and serve a target for the development of new therapies.

Trefoil factors in saliva and gingival tissues of patients with chronic periodontitis.

BACKGROUND: Trefoil factors (TFFs) are secreted molecules that are involved in cytoprotection against tissue damage and the immune response. TFFs have been detected in saliva and oral tissues, but their clinical significance has never been investigated in patients with chronic periodontitis. The objective of this study is to determine whether TFF expression in saliva and gingival tissues is associated with periodontal pathology. METHODS: Saliva and gingival tissue samples were collected from 25 non-periodontitis individuals and 25 patients with chronic periodontitis (CP). Enzyme-linked immunosorbent assay and immunohistochemical methods were used to evaluate the expression of TFFs (TFF1, TFF2, and TFF3) in saliva and gingival tissues, respectively. Periodontopathic bacteria were quantified by real-time polymerase chain reaction. RESULTS: Reduced salivary TFF1 and TFF3 concentrations were observed in patients with CP (P = 0.003 and P <0.001, respectively). Decreased TFF3 expression in gingival tissues of patients with CP was demonstrated (P = 0.041). Levels of salivary TFF3 concentrations were negatively correlated with periodontal pathology and number of Porphyromonas gingivalis and Tannerella forsythia (formerly known as Bacteroides forsythus). CONCLUSIONS: Altered expression of TFFs in saliva and gingival tissues was detected in patients with CP. The results suggest that TFF3 may be involved in the pathogenesis of periodontal disease.

Effect of Curcuma comosa and estradiol on the spatial memory and hippocampal estrogen receptor in the post-training ovariectomized rats.

Curcuma comosa was reported to have uterotrophic effects in ovariectomized (OVX) rats. The present study aims to investigate the plant estrogenic-like effects of C. comosa hexane extract on spatial memory and estrogen receptors (ER) in the hippocampus. Female rats received 5 days' Morris water maze training before the ovariectomy, followed by the daily intraperitoneal administration of C. comosa hexane extract at a dose of 250 and 500 mg/kg body weight for 4 weeks. Estradiol (10 µg/kg body weight) was used as the positive control. OVX significantly increased the latency of the animal to the platform, while estradiol and both doses of C. comosa hexane extract reversed the memory impairments. By using the reverse transcription polymerase chain reaction technique, C. comosa hexane extract was shown to produce a selective increase of the ER alpha subtype but not beta subtype in hippocampus, while estradiol increased both of them. C. comosa did not affect spatial memory, learning ability, and ER mRNA levels in the sham OVX rats. In conclusion, C. comosa hexane extract improved the spatial memory of the OVX rats, which was similar to estrogen. The influence on the memory changes might be selectively related to ER alpha in the hippocampus.

Relationship between periodontitis and diabetes - importance of a clinical study to prove the vicious cycle.

Periodontitis is regarded to have a close relationship to diabetes mellitus. In Japan, some cohort studies have indicated that there is a significant positive association of obesity plus metabolic syndrome with periodontal diseases. As such, an increasing number of studies suggest a relationship between periodontitis and diabetes, most of which are epidemiologic association. In this review, we have summarized the possible evidence of a relationship between periodontitis and diabetes. To date, little evidence has been reported to indicate that diabetes and/or glucose intolerance has in fact had a significant cause-effect relationship with periodontal disease. In this regard, it is important to directly uncover the relation, i.e., to prove the effect of therapeutic approaches to periodontitis upon mitigation of glycemic control in diabetic patients, which would be a direct evidence of its causal nature. Therefore a study should be undertaken to this effect.

Alanine aminopeptidase and dipeptidyl peptidase IV in saliva of chronic periodontitis patients.

BACKGROUND: Alanine aminopeptidase (ALAP) and dipeptidyl peptidase IV (DPPIV) are ectopeptidases that play a role in collagen degradation and are thought to be involved in the destruction of periodontal tissue. This study compared the activities of salivary ALAP and DPPIV in patients with periodontitis and periodontally healthy subjects. The correlations of enzyme activities with clinical variables and the presence of Porphyromonas gingivalis were also evaluated. METHODS: Whole saliva was collected from 30 periodontally healthy subjects, 30 localized chronic periodontitis (LCP) patients, and 30 generalized chronic periodontitis (GCP) patients to determine the activities of ALAP and DPPIV. The presence of P. gingivalis in subgingival plaque was detected by polymerase chain reaction. Periodontal clinical assessments included probing depth, clinical attachment level, and bleeding on probing. RESULTS: The activities of DPPIV in the LCP and GCP groups were not significantly different from one another, but both groups had significantly higher enzyme activities than the periodontally healthy group (P = 0.001). DPPIV activity was positively correlated with all clinical parameters and the prevalence of P. gingivalis. The ALAP activities were not significantly different among the three study groups. There was no significant correlation of ALAP activity with any of the clinical and bacterial parameters. CONCLUSION: DPPIV, but not ALAP, activity is associated with periodontitis and the presence of P. gingivalis.

Association between periodontitis and anti-cardiolipin antibodies in Buerger disease.

AIM: Anti-cardiolipin (CL) antibodies can be induced in Buerger disease (BD), an inflammatory occlusive disorder affecting peripheral blood vessels, in response to bacteria bearing homology to the TLRVYK peptide of a phospholipid-binding plasma protein beta-2-glycoprotein I. TLRVYK homologies are present in Porphyromonas gingivalis (TLRIYT) and Treponema denticola (TLALYK). This study investigated the association between periodontal infection and anti-CL antibodies in BD patients. MATERIAL AND METHODS: Periodontal conditions were examined in 19 BD patients and 25 systemically healthy control subjects. All subjects were heavy smokers. Serum anti-CL, anti-TLRVYK, anti-TLRIYT, and anti-TLALYK antibodies were assessed using the enzyme-linked immunosorbent assay. RESULTS: BD patients had a significantly higher prevalence of periodontitis, more severe periodontal destruction and increased titres of serum anti-CL, anti-TLRVYK, anti-TLRIYT, and anti-TLALYK antibodies compared with healthy subjects. The levels of anti-CL antibodies positively correlated with those of the three anti-peptide antibodies. Anti-CL antibody titres were significantly associated with the percentage of sites with clinical attachment level >or=4 mm in BD patients. CONCLUSION: Elevated anti-CL antibody levels were associated with periodontal destruction in BD patients. Periodontopathic bacteria may serve as exogenous antigens that stimulate the anti-CL antibody production through molecular mimicry between the bacterial peptides and a host plasma protein.

Anti-inflammatory effect of Streblus asper leaf extract in rats and its modulation on inflammation-associated genes expression in RAW 264.7 macrophage cells.

AIM OF THIS STUDY: Streblus asper is a medicinal plant from Thailand used in folk medicine for the treatment of several inflammatory diseases. In this study, we investigated the anti-inflammatory effect of Streblus asper leaf ethanolic extract (SAE). MATERIALS AND METHODS: The experimental carrageenan-induced paw edema in rats was performed in which the SAE at doses of 125, 250, 500 mg/kg body weight was intraperitoneally administered to the rats. Then, reverse transcriptive polymerase chain reaction (RT-PCR) technique was also performed to determine the effect of SAE on the expression of inflammation-associated genes in RAW 264.7 macrophage cells stimulated with lipopolysaccharide (LPS). RESULTS: The SAE at all given doses caused a significant dose-dependent inhibition of edema (p<0.05). Moreover, the significant and dose-dependent LPS-induced cyclooxygenase (COX)-2 and inducible nitric oxide synthase (iNOS) mRNA expressions were demonstrated in RAW 264.7 cells treated with SAE. The inhibition is selective, since COX-1 mRNA expression did not change in the presence of SAE. CONCLUSION: The results of this study are the first scientific evidence on the molecular effects of Streblus asper as a potential anti-inflammatory agent, which supports the fact that the plant is employed in traditional remedies.

C-reactive protein associated with periodontitis in a Thai population.

AIM: C-reactive protein (CRP) has been implicated as a possible mediator of the association between periodontitis and several systemic diseases. Previous studies suggest an association between increased CRP levels and periodontitis predominantly in Caucasians. This study evaluated the associations of chronic periodontitis and Porphyromonas gingivalis with CRP in systemically healthy Thai adults. MATERIAL AND METHODS: Serum high-sensitivity CRP was measured in 21 generalized periodontitis, 62 localized periodontitis, and 38 periodontally healthy control subjects. P. gingivalis in subgingival plaque samples was analyzed by polymerase chain reaction. RESULTS: Overall, these subjects had a median CRP level lower than that reported in the western populations. Subjects with generalized periodontitis and localized periodontitis had higher median CRP levels than controls (1.78 and 0.65 mg/l versus 0.25 mg/l, p<0.001). Multivariate linear regression showed that log CRP levels were increased in subjects with generalized periodontitis (p<0.01) and localized periodontitis (p=0.03) compared with the controls, adjusted for age, body mass index and smoking. Presence of P. gingivalis was also independently associated with elevated log CRP levels (p<0.001). CONCLUSION: Periodontitis and subgingival P. gingivalis are associated with increased CRP levels. These findings suggest that periodontal infection may contribute to systemic inflammatory burden in otherwise healthy individuals.

Differential response of cytokines induced by Leptospira interrogans, serogroup Pomona, serovar Pomona, in mouse and human cell lines.

Leptospira interrogans, the causative agent of leptospirosis, is an important zoonotic bacterium. The mechanisms and roles of cytokine induction in both humans and animals remain unclear. Therefore, the IFN-gamma, IL-6, IL-10 and IL-12 levels were measured by enzyme-linked immunosorbent assay (ELISA) in human THP-1 and mouse RAW 264.7 monocyte cell lines following stimulation with heat-killed Leptospira interrogans serogroup Pomona serovar Pomona, L. biflexa, E. coli or Salmonella group B. The production of IL-6 and IL-12 were higher and rose more rapidly in the RAW 264.7 cells with all bacteria. The IL-10 was not detected in the RAW 264.7 cells when induced by leptospires. The IFN-gamma level in human peripheral blood mononuclear cells (PBMCs) induced by leptospires was also significantly lower than with other bacteria. When IL-10 and IL-12 mRNA expressions were detected in hamster's spleen, their patterns were similar to what was observed in THP-1 in that IL-12 was only slightly increased while IL-10 expression was high. Moreover, the IFN-gamma expression could not be detected in hamsters. The more potent cytokine responses in the RAW 264.7 cells may indirectly reflect the disease outcome in mice which render them to be a good reservoir of leptospirosis. Whether these cytokines have contributed to immunoprotection during the L. interrogans infection remains to be further investigated.

RANKL upregulation associated with periodontitis and Porphyromonas gingivalis.

BACKGROUND: Receptor activator of nuclear factor-kappa B (NF-kappaB) ligand (RANKL) and osteoprotegerin (OPG) are critical for homeostatic control of osteoclast activity, suggesting their vital roles in the progression of bone loss in periodontitis. In this study, the expression of RANKL and OPG mRNA and the relationship between these factors and periodontopathic bacteria in periodontal tissue were studied. METHODS: Gingival tissue and subgingival plaque samples were collected from 15 patients with chronic periodontitis and 15 periodontally healthy subjects. RNA was extracted from the tissue and subjected to reverse transcription-polymerase chain reaction (RT-PCR) using primers specific for RANKL or OPG. Beta-actin was amplified as a control to ensure equal loading. The intensity of RT-PCR products was analyzed by a densitometer in proportion to the intensity of beta-actin. The numbers of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans were determined by quantitative real-time PCR. RESULTS: Our results showed increased levels of RANKL mRNA in chronic periodontitis tissues. The RANKL/OPG expression ratio was significantly higher in the periodontitis group compared to the healthy control group (P = 0.001). Interestingly, the expression of RANKL (r = 0.64; P <0.001), but not OPG (r = -0.24; P = 0.20), was significantly correlated with increased numbers of P. gingivalis. A. actinomycetemcomitans was detected in only 6.7% of all sites. CONCLUSIONS: Chronic periodontitis was associated with RANKL mRNA upregulation and increased RANKL/OPG mRNA expression ratio. In addition, our data showed for the first time to our knowledge an association between upregulated RANKL levels and the number of P. gingivalis in clinically obtained periodontal tissues.

TRAF6 activation of PI 3-kinase-dependent cytoskeletal changes is cooperative with Ras and is mediated by an interaction with cytoplasmic Src.

Interleukin 1 (IL-1) has been implicated in the reorganization of the actin cytoskeleton. An expression vector encoding a PKB/Akt pleckstrin-homology domain fused to a fluorescent protein was used to detect phosphoinositide 3-kinase (PI 3-kinase) products. It was observed that PI 3-kinase was activated either by treatment with IL-1 or by expression of either TRAF6, Src, MyD88 or dominant-positive PI 3-kinase, and resulted in the formation of long filopodia-like cellular protrusions that appeared to branch at membrane sites consisting of clusters of phosphoinositide. This depended upon a TRAF6 polyproline motif and Src catalytic activity, and was blocked by inhibitors of PI 3-kinase, Src and Ras. Using both conventional and split fluorescent protein probes fused to expressed TRAF6 and Src in living cells, the polyproline sequence of TRAF6 and the Src-homology 3 (SH3) domain of Src were shown to be required for interaction between these two proteins. Interaction occurred within the cytoplasm, and not at either the cell membrane or cytoplasmic sequestosomes. In addition, co-transfection of vectors expressing fluorescent-protein-fused TRAF6 and non-fluorescent MyD88, IRAK1 and IRAK2 revealed an inverse correlation between increased sequestosome formation and activation of both PI 3-kinase and NF-kappaB. Although a key factor in TRAF6-dependent activation of PI 3-kinase, ectopic expression of Src was insufficient for NF-kappaB activation and, in contrast to NF-kappaB, was not inhibited by IRAK2.

Conserved ETS domain arginines mediate DNA binding, nuclear localization, and a novel mode of bZIP interaction.

The DNA-binding ETS transcription factor Spi-1/PU.1 is of central importance in determining the myeloid-erythroid developmental switch and is required for monocyte and osteoclast differentiation. Many monocyte genes are dependent upon this factor, including the gene that codes for interleukin-1beta. It has long been known that the conserved ETS DNA-binding domain of Spi-1/PU.1 functionally cooperates via direct association with a diverse collection of DNA-binding proteins, including members of the basic leucine zipper domain (bZIP) family. However, the molecular basis for this interaction has long been elusive. Using a combination of approaches, we have mapped a single residue on the surface of the ETS domain critical for protein tethering by the C/EBPbeta carboxyl-terminal bZIP domain. This residue is also important for nuclear localization and DNA binding. In addition, dependence upon the leucine zipper suggests a novel mode for both protein-DNA interaction and functional cooperativity.

A novel mutation of the cathepsin C gene in a thai family with Papillon-Lefevre syndrome.

BACKGROUND: Papillon-Lefevre syndrome (PLS) is a rare autosomal recessive disorder characterized by palmar- plantar hyperkeratosis and rapid periodontal destruction of both primary and permanent dentitions. It has been shown that the disease is caused by cathepsin C gene (CTSC) mutation leading to the deficiency of cathepsin C enzymatic activity. This study demonstrates the clinical manifestations and CTSC mutational and enzymatic activity analyses in a 5-year-old Thai male PLS patient and his parents. METHODS: Peripheral blood samples were obtained for genomic DNA isolation. All exons of the CTSC gene were amplified by polymerase chain reaction (PCR) using specific primers. Mutations were identified by DNA sequencing. Verification of the mutation was performed by digestion of PCR products by restriction endonucleases. The cathepsin C enzymatic activity was determined using the synthetic substrate glycyl- L-arginine-7-amino-4-methylcoumarin. RESULTS: The patient demonstrated classical characteristics of PLS, including hyperkeratotic skin lesions. By the age of 5, all of his primary teeth were extracted due to severe periodontal infection. The parents had no physical abnormalities. The periodontal examination revealed localized mild periodontal destruction. Sequence analysis showed a nucleotide change at position 90 from C >A (c.90C >A) which resulted in a change from cysteine residue to a premature stop codon at the amino acid position 30 in the exon 1. The HpyCH4V digestion revealed that the patient was homozygous, whereas both the father and mother were heterozygous carriers of this mutation. The cathepsin C activity was reduced in the patient's mother, and the activity in the patient was almost completely lost. CONCLUSIONS: This is the first study to demonstrate a CTSC gene mutation in a Thai family with PLS. The identified mutation is novel and potentially leads to the drastic reduction of the cathepsin C enzymatic activity. This suggests that the mutation is pathogenetic, causing the PLS. Mutational analysis in more members of the family is warranted to identify whether the mutation is inherited from a common ancestor.