Effect of Botulinum Toxin Injection on EMG Activity and Bite Force in Masticatory Muscle Disorder: A Randomized Clinical Trial.

Botulinum toxin type A (BoNT-A) is increasingly used in treating masticatory muscle pain disorder; however, safe doses and reinjection intervals still need to be established. The purpose of this randomized clinical trial was to evaluate the degree and duration of the impairment of masticatory muscle performance. Fifty-seven subjects were randomly divided into two groups: one of which received BoNT-A first (n = 28) while the other received saline first (n = 29), with the cross-over being in week 16, and a total follow-up period of 32 weeks. A total dose of 50 U of BoNT-A was injected in the masseter and temporal muscles bilaterally. Electromyographic (EMG) activity and bite forces were assessed. A significant reduction in EMG activity was observed up to week 18 (p ≤ 001), with total recovery at week 33. A significant reduction in maximum bite force was observed up to week 11 (p ≤ 005), with total recovery at week 25. In conclusion, when treating masticatory muscle pain disorder with 50 U of BoNT-A, a reinjection interval of 33 weeks can be considered safe since the recovery of muscle function occurs by that time.

Comprehensive evaluation of botulinum toxin treatment outcomes of a patient with persistent myofascial orofacial pain.

The outcome evaluation method presented in this case study, including Axes I and II findings combined with the results of quantitative bite force and EMG measurements, provides a good tool for proper evaluation of the effect of BoNT-A on patients with myofascial orofacial pain and changes in jaw muscle function.

Common Matrix Metalloproteinases (MMP-8, -9, -25, and -26) Cannot Explain Dentigerous Cyst Expansion.

OBJECTIVE: Mechanisms of the dentigerous cyst formation from the normal eruption follicle is unknown but disturbances in the proteolytic activity have been suspected, since the growth of these cysts is accompanied by local bone destruction. The aim of the present study was to evaluate the expression of matrix metalloproteinases (MMP) in human dental dentigerous cysts and healthy dental follicles. MATERIALS AND METHODS: We studied 10 patients with dentigerous cysts and 10 healthy dental follicles from the lower jaw in respect to their immunoexpression of MMPs -8, -9, -25, and -26 and tissue inhibitor of metalloproteinases -1 (TIMP-1). RESULTS: MMP-8 was expressed slightly more in cyst epithelium than in odontogenic epithelium of healthy controls dental follicle but the difference lacked statistical difference. Other MMPs and TIMP-1 did not differ regarding the studied specimens. CONCLUSION: Differences in MMP expression cannot solely explain the cyst expansion suggesting the potential involvement of other osteolytic mechanisms.

Delayed diagnosis of oral cancer in Iran: challenge for prevention.

PURPOSE: The aim of this study was to investigate the diagnostic delay and its determinants among oral cancer patients in Tehran, Iran. MATERIALS AND METHODS: This study was conducted between September 2004 and September 2006 in three university hospitals, and included 100 consecutive patients with primary oral squamous cell carcinoma (international classification of disease, ICD-10 sites C01 to C06). Data were obtained through questionnaire interviews and medical records of the patients were reviewed to obtain information on the date of diagnosis, primary tumour site and the stage of the tumour at the time of diagnosis. Statistical analysis was performed by t test, ANOVA and logistic regression. RESULTS: The mean diagnostic delay was 7.2 months (SD 7.5, range 1 to 36 and median 4). The most important determinants of longer diagnostic delay were being single (OR = 4.8; 95% CI = 1.5 to 14.8; P < 0.05) and being at advanced tumour stages (OR = 5.3; 95% CI = 1.8 to 15.6; P < 0.01). The mean patient and professional delays were 5.3 months (SD 6.1 and median 2) and 2.1 months (SD 2.1 and median 1), respectively. Patients at advanced tumour stages were more likely to have longer patient and professional delays than those at early stages (OR = 5.6; 95% CI = 1.8 to 17.3 and OR = 3.4; 95% CI = 1.2 to 9.4, respectively; P < 0.05). Living alone was also a determinant of longer patient and professional delays (OR = 7.1; 95% CI = 2.0 to 24.7, OR = 3.5; 95% CI = 1.2 to 10.3, respectively; P < 0.05). CONCLUSIONS: Developing preventive programmes that focus on the enhancement of public and professional awareness about oral cancer is essential to promote earlier diagnosis in Iran.

Survival after lip cancer diagnosis.

The purpose of this study was to analyze the 5-year survival rates of 82 patients with lip cancer attending 5 university hospitals during 1999-2003 in Tehran, Iran. We used information from patient records, telephone calls, and death register files of the Iran Ministry of Health to ascertain the patients' vital status. Associations between survival and the variables of sex, age, stage of the tumor at the time of diagnosis, treatment modality, and tumor histopathologic type were analyzed with Kaplan-Meier, log-rank, and Cox regression methods. Of all patients, 70 (85%) were men, with a median age of 62 years (mean, 58.6 years [SD, 15 years]; range, 27-85 years) at the time of diagnosis. The median follow-up time of the patients was 57 months (mean, 56.4 months [SD, 28 months]; range, 0-112 months). The 1- to 5-year overall survival rate was 91% to 62%. The tumor stage at the time of diagnosis and the treatment modality were associated with survival (P < 0.05) in both univariate and multivariable analyses. Patients who underwent surgery and had lower stage tumors at the time of diagnosis showed higher survival rates. No differences in patient survival were found regarding sex, age, and histopathologic type of tumors. These findings indicate that although lip tumors are curable, early detection, diagnosis, and treatment lead to even higher rates of survival. Importance of the early detection of lip cancer should be emphasized in all health care and cancer prevention campaigns directed to the public and professionals.

Survival after diagnosis of cancer of the oral cavity.

In this retrospective study we analysed the survival in 470 patients with oral cancer. Patients who attended five university hospitals in Tehran, Iran, during the period 1996-2002 were included. Data were obtained from a combination of sources including patients' records, telephone calls, and deaths registered by the Ministry of Health. Survival curves were generated using Kaplan-Maier curves. Univariate and multivariate analyses of the relations between survival and age, sex, site of primary tumour, stage, and histopathological type were made using the log-rank test and Cox's regression analysis. Sex and age were not associated with survival. Treatment and stage of tumour at the time of diagnosis were related to survival. The overall survival rates were higher in patients with stages I or II cancer than those in stages III (OR=2.8, 95% CI=1.8 to 4.4) or IV (OR=4.6, 95% CI=3.1 to 6.8) at the time of diagnosis. Patients treated with radiotherapy had lower survival than those who had been operated on and had radiotherapy or operation alone (OR=2.8, 95% CI=1.7 to 4.5). There was no difference in survival depending on the histological type of tumour. To achieve higher survival, early detection and diagnosis of oral cancers should be emphasised in oral health programmes to improve public awareness and preventive activities among dentists in Iran.

Malignant oral tumors in iran: ten-year analysis on patient and tumor characteristics of 1042 patients in Tehran.

This study analyzed characteristics of oral cancer patients from Tehran, Iran, and their tumors. Data came from the patient records of 30 major hospitals in Tehran. Patients (n = 1042), diagnosed with invasive oral cancer in 1993-2003, were classified by primary tumor site according to ICD-10 (C00-C10). Data were analyzed separately for lip, oral cavity and salivary gland tumors. Statistical evaluation included chi and t-test. Of all cases, 59% were male. Age for all cases ranged from 6-103 years, mean age was 58.8 years (SD 16; median 62); 89% were older than 40. Tumor site breakdown was 65% oral cavity, 21% major salivary glands and 14% lip. A clear gender difference (P < 0.001) appeared regarding the primary tumor sites: women dominated in oral cavity cancers and men in lip cancers. The most common cancer site was the tongue (32%), accounting for 50% of the oral cavity cancers. Histologically, 88% of all oral cavity and lip cancers were squamous cell carcinomas, 10% of those were in age /= age 65. At the time of diagnosis, 59% of oral cavity cancers and 29% of lip cancers were at stage III or IV (P < 0.001). The results emphasize an urgent need for a national program focusing on early detection of oral cancers, including educational information addressed to oral health professionals.

Immunohistochemical evaluation of inflammatory mediators in failing implants.

It was hypothesized that peri-implant tissue around loosening dental implants may contain cytokines with a potential to regulate osteoclasts. Peri-implant and/or gingival samples from loosened implants, chronic periodontitis (CP), and normal controls (n = 10 samples in each group) were analyzed using immunohistochemical staining to observe tumor necrosis factor alpha (TNF-alpha), interleukin 1-alpha (IL-1alpha), IL-6, platelet-derived growth factor A (PDGF-A), and transforming growth factor alpha (TGF-alpha). These cytokines were found in foreign-body giant cells, macrophages, fibroblasts, and epithelial cells. TNF-alpha, IL-1alpha, and IL-6 were increased (P < .05; unpaired t test) in peri-implantitis and CP, whereas PDGF-A and TGF-alpha were not. In conclusion, cytokines with a potential to activate osteoclasts were found in both peri-implantitis and CP, but the cytokine profiles differed in that IL-1alpha was the most prevalent cytokine in the former and TNF-alpha was the most common in the latter. These cytokines may contribute to peri-implant bone loss/loosening by stimulating formation and activity of osteoclasts and might be an amenable target for local therapies with cytokine modulators.

Levels and molecular forms of MMP-7 (matrilysin-1) and MMP-8 (collagenase-2) in diseased human peri-implant sulcular fluid.

OBJECTIVES: Matrix metalloproteinases (MMPs) play crucial role in various tissue destructive inflammatory processes by degrading almost all peri-cellular and basement membrane components. MMP-8 (collagenase-2) is the major MMP in periodontitis. MMP-7 (matrilysin-1), in addition to its ability to degrade matrix and basement membrane components, activates other latent pro-MMPs and defensins, host cell-derived antimicrobial cryptidins. The aim of the present study was to characterize the relationship, levels and molecular forms of MMP-8 and MMP-7 in diseased peri-implant sulcular fluid (PISF). MATERIALS AND METHODS: Seventy-two human dental implant fluid samples were collected with filter paper strips from peri-implant sulci from healthy and untreated diseased implant sites. Gingival index (GI) and/or bone resorption (BR) were also recorded. Western immunoblot method with polyclonal anti-human-MMP-8 and monoclonal anti-human-MMP-7 antibodies was used, and immunoreactivities were quantified with computer scanning program. The effects of MMP inhibitors (doxycycline, chemically modified tetracycline-3, clodronate, CTT-peptide and marimastat) were studied on the activity of recombinant human matrilysin-1 (MMP-7) using beta-casein degradation assay. RESULTS: The levels of active forms of MMP-8 and MMP-7 were significantly elevated in diseased PISF in relation to healthy PISF. Furthermore, MMP-8 and MMP-7 levels correlated significantly to each other and GI. MMP-8 was present not only as bands corresponding to 75-kDa polymorphonuclear leukocyte (PMN) -type pro- and 65-kDa active forms, but also as 55-kDa non-PMN-type pro- and 45-kDa active forms. Immunoreactivities > 80 kDa most likely represented dimeric and/or inhibitor-bound MMP-8 complexes and the low molecular weight (< 30 kDa) species were apparently degraded fragments. In diseased PISF, 19-21-kDa active MMP-7 and 28-30-kDa pro-MMP-7 species were detected, and the active 19-21-kDa forms of MMP-7 predominated in diseased PISF. Doxycycline (50 micro m and 250 micro m), chemically modified non-antimicrobial tetracycline (CMT-3) (50 micro m and 100 micro m), clodronate (a bisphosphonate, 20 micro m and 500 micro m) and the cyclic CTT (CTTHWGFTLC)-peptide (125 micro m and 250 micro m), all known broad-spectrum or selective MMP-inhibitors, did not inhibit the activity of human recombinant MMP-7; only marimastat (1 micro m and 5 micro m) inhibited MMP-7. DISCUSSION: Increased immunoreactivities of the active MMP-8 and MMP-7 species in PISF from diseased peri-implantitis lesions eventually reflect the stage and course of peri-implantitis; MMP-7 may potentially act as MMP-8 and defensin activator in diseased PISF. CONCLUSION: The elevated levels of MMP-8 and matrilysin-1/MMP-7 were identified in active forms in diseased PISF, but MMP-7 was less prominent. MMP inhibitors, potential future tissue protective drugs, seemingly do not interfere with the defensive antibacterial action of MMP-7.

Inhibition of matrix metalloproteinase-14 in osteosarcoma cells by clodronate.

BACKGROUND: Bisphosphonates reduce the bone metastasis formation and angiogenesis but the exact molecular mechanisms involved are unclear. Progelatinase A (proMMP-2; 78 KDa) is activated up during the tumor spread and metastasis by a cell surface-associated matrix metalloproteinase (membrane-type matrix metalloproteinase [MT1-MMP] or MMP-14). MATERIAL AND METHODS: We evaluated the effects of a bisphosphonate (clodronate) on MT1-MMP mRNA expression and protein production, catalytic activity and proteolytic activation of proMMP-2 by cultured human MG-63 osteosarcoma cells. RESULTS: Clodronate, at therapeutically attainable noncytotoxic concentrations, dose-dependently inhibited phorbol myristic acetate (PMA)-induced proteolytic activation of proMMP-2 by human MG-63 osteosarcoma cells. Clodronate also downregulated the PMA-induced expression of MT1-MMP mRNA and protein production in human MG-63 osteosarcoma cells, as evidenced by Northern analysis and fluorescent immunohistochemistry. Furthermore, clodronate inhibited directly and dose-dependently MT1-MMP activity, and the MT1-MMP inhibition by clodronate was reduced in the presence of an increased (5 mM) Ca(2+) concentrations when compared to physiological (1 mM) Ca(2+) concentrations. CONCLUSION: We conclude that (1) the extracellular/cell-associated mechanism of bisphosphonate involves inhibition of MT1-MMP catalytic activity eventually by chelation, and that (2) intracellular mechanism involves downregulation of induced MT1-MMP mRNA and protein expression. The inhibition and downregulation of MT1-MMP by clodronate can be related to their ability to reduce MG-63 osteosarcoma cell invasion and spread. These findings may, at least in part, explain at molecular level the antitumor and antibone resorption activities of clodronate observed in clinical studies.

Laminin-5 gamma2-chain and collagenase-2 (MMP-8) in human peri-implant sulcular fluid.

Laminin-5 (LN-5) is an important epithelial cell-derived structural and adhesive component in hemidesmosomes and basement membranes (BM). In peri-implant tissue, gingival BM underlies the junctional epithelium (JE) and reflects the peri-implant health. Matrix metalloproteinase-8 (MMP-8 or collagenase-2) is one of the key mediators of periodontal tissue destruction. Western immunoblotting with image analysis was used to quantitate the molecular forms of LN-5 gamma2-chain and MMP-8 in peri-implant sulcular fluid (PISF) from healthy and diseased implants. These observations were related to the recorded gingival (GI) and bone resorption (BR) indices of the studied sites. Altogether, 72 PISF samples from osseointegrated dental implants were examined. Significantly elevated levels of fragmented LN-5 gamma2-chain species (45 and 70 kDa) and MMP-8 immunoreactivities were observed in diseased PISF in relation to healthy PISF. The elevated levels of both LN-5 gamma2-chain 45 and 70 kDa fragments and MMP-8 in diseased PISF from peri-mucositis (BR = 0) and peri-implantitis (BR >/= 1) lesions strongly correlated with elevated GI. Low levels - almost comparable to those seen in healthy control PISF - were seen in PISF from peri-implantitis lesions (BR >/= 1) with no GI. Activation of 75 kDa neutrophil (PMN)-type proMMP-8 to 10 kDa lower-molecular-size active forms was especially detected in PISF from peri-implantitis with elevated GI. These cross-sectional findings indicate that elevated MMP-8 and LN-5 gamma2-chain fragment levels in PISF can reflect the active phase of the inflammatory peri-implant disease. Longitudinal studies are required to assess their use, either alone or in combination as molecular biochemical PISF markers, to predict the risk of progression of peri-implantitis, as well as to monitor the impact of treatment of the disease.

Laminin-5 gamma 2 chain is colocalized with gelatinase-A (MMP-2) and collagenase-3 (MMP-13) in odontogenic keratocysts.

BACKGROUND: Odontogenic keratocyst (KC) differs from other epithelial odontogenic cysts in regard to increased epithelial proliferation and a strong tendency to recur. Laminin-5 (Ln-5) is an epithelial anchoring filament component, which after modulation by certain matrix metalloproteinases (MMPs), like MMP-2 and MMP-13, induces epithelial cell migration. METHODS: Using in situ hybridization and immunohistochemistry, we studied the Ln-5 gamma-2 chain expression related to the expression of MMP-2, -8, and -13 in different odontogenic cysts, including radicular cysts (RC; n = 11), follicular cysts (FC; n = 11), and odontogenic keratocysts (KC; n = 16). RESULTS: Ln-5 mRNA was present in all cysts examined, while less than half of KCs and RCs (33 and 40%, respectively) demonstrated MMP-2 mRNA. MMP-13 mRNA was present in all KC samples. Ln-5 protein was located as a continuous ribbon in BM zone of all KCs, and MMP-2 and MMP-13 immunoreactivities colocated significantly with Ln-5 in that area. MMP-8 was expressed by stromal macrophages and epithelial goblet cells, but never located in BM zone. CONCLUSIONS: Our results indicate that the colocalization of Ln-5 with MMP-2 or MMP-13, but not with MMP-8, in BM zone of KCs, may be related to special characteristics of KC.

Gelatinase B is associated with peri-implant bone loss.

The aim of this study was to clear whether gelatinase B is associated with peri-implant bone loss (PBL). Peri-implant sulcus fluid was collected from 46 implant sites in 12 patients. These sites were also characterized using modified Gingival Index (mGI). Activated and total gelatinase B levels, measured using a modified urokinase assay, showed correlation with PBL (n = 46, Spearman's rank correlation test). Activated and total gelatinase B values were significantly higher in PBL > 3 mm group (n = 6) compared to PBL < 1 mm (n = 29) and 1 < PBL < 3 mm (n = 11) groups (rank sum test). Activated gelatinase B level in mGI > 0.5 group (n = 24) was clearly higher compared to mGI = 0 (n = 13) and < or = 0.5 (n = 9) groups (Rank sum test). We conclude that gelatinase B is associated with PBL. Activation of gelatinase B together with elevated mGI eventually reflect active phases of peri-implantitis and may prove to be diagnostically useful.

Bisphosphonates inhibit stromelysin-1 (MMP-3), matrix metalloelastase (MMP-12), collagenase-3 (MMP-13) and enamelysin (MMP-20), but not urokinase-type plasminogen activator, and diminish invasion and migration of human malignant and endothelial cell lines.

Bisphosphonates (clodronate, alendronate, pamidronate and zoledronate) at therapeutically attainable non-cytotoxic concentrations inhibited MMP-3, -12, -13 and -20 as well as MMP-1, -2, -8 and -9, but not urokinase-type plasminogen activator (uPA), a serine proteinase and a pro-MMP activator. Dose-dependent inhibition was shown by three independent MMP assays. The inhibition was reduced in the presence of an increased concentration of Ca(2+) when compared to physiologic Ca(2+) concentration. Alendronate inhibited the in vitro invasion (Matrigel) of human HT1080 fibrosarcoma and C8161 melanoma cells, and the random migration of these malignant and endothelial cell lines capable of expressing MMPs and uPA. The concentration of alendronate required to inhibit 50% of the activity (IC(50)=40-70 microM) of MMPs corresponded to the IC(50) of down-regulation of in vitro invasion and migration. The ability of bisphosphonates to down-regulate the in vitro invasion and random migration was comparable or slightly better in relation to the selective gelatinase inhibitor CTTHWGFTLC peptide. Alendronate but not CTTHWGFTLC peptide promoted the adhesion of HT1080 fibrosarcoma and C8161 melanoma cell lines on fibronectin. Bisphosphonates are broad-spectrum MMP inhibitors and this inhibition involves cation chelation. Bisphosphonates further exert antimetastatic, anti-invasive and cell adhesion-promoting properties, which may prevent metastases not only into hard tissues but also to soft tissues.