Unique Raman Spectroscopic Fingerprints of B-Cell Non-Hodgkin Lymphoma: Implications for Diagnosis, Prognosis and New Therapies.

OBJECTIVE: Raman spectroscopy is a non-invasive laser-based technique that identifies molecular chemical composition of tissues and cells. The objective of the work was to demonstrate that unique Raman spectroscopic fingerprints of B-cell non-Hodgkin lymphoma cells could be distinguished from normal B-cells. METHODS: Normal B-cells and B-cell non-Hodgkin lymphoma cells were mounted on aluminum slides and analyzed by Raman spectroscopy using Asymmetric Least Squares and Principal Component Analysis. RESULTS: Clustering by Principal Component Analysis differentiated normal B-cells from B-cell non-Hodgkin lymphoma cells as well as between the different B-cell non-Hodgkin lymphoma cell types. CONCLUSIONS: Raman spectroscopy technology provided a different paradigm in analyzing tumor cells which could be used for diagnosis as well as contribute new information on unique characteristics of cancer cells to understand pathogenesis and potential novel treatments.

EZH2-mediated upregulation of ROS1 oncogene promotes oral cancer metastasis.

Current anti-epidermal growth factor receptor (EGFR) therapy for oral cancer does not provide satisfactory efficacy due to drug resistance or reduced EGFR level. As an alternative candidate target for therapy, here we identified an oncogene, ROS1, as an important driver for oral squamous cell carcinoma (OSCC) metastasis. Among tumors from 188 oral cancer patients, upregulated ROS1 expression strongly correlated with metastasis to lung and lymph nodes. Mechanistic studies uncover that the activated ROS1 results from highly expressed ROS1 gene instead of gene rearrangement, a phenomenon distinct from other cancers. Our data further reveal a novel mechanism that reduced histone methyltransferase EZH2 leads to a lower trimethylation of histone H3 lysine 27 suppressive modification, relaxes chromatin, and promotes the accessibility of the transcription factor STAT1 to the enhancer and the intron regions of ROS1 target genes, CXCL1 and GLI1, for upregulating their expressions. Down-regulation of ROS1 in highly invasive OSCC cells, nevertheless, reduces cell proliferation and inhibits metastasis to lung in the tail-vein injection and the oral cavity xenograft models. Our findings highlight ROS1 as a candidate biomarker and therapeutic target for OSCC. Finally, we demonstrate that co-targeting of ROS1 and EGFR could potentially offer an effective oral cancer therapy.

Polymorphisms in Kallikrein7 and 10 genes and oral cancer risks in Taiwan betel quid chewers and smokers.

OBJECTIVES: We investigated the association between mRNA levels, polymorphisms of Kallikrein7 (KLK7) and Kallikrein10 (KLK10), and the development of oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: We recruited 217 OSCC patients and 138 healthy controls. All were men, betel quid chewers, cigarette smokers, and Minnan ethnicity. Genotyping was performed using a TaqMan probe genotyping assay. Gene expression levels were determined using real-time polymerase chain reactions (PCRs) for 20 pairs of cancerous and non-cancerous tissues. RESULTS: Kallikrein10 rs3745535G>T polymorphisms were significantly associated with OSCC development [adjusted OR (AOR) = 1.62, 95% CI = 1.02-2.59], but KLK7 polymorphisms were not. The KLK7 rs10581213(wt/ins + ins/ins) genotypes were significantly associated with early-stage cancer (AOR = 0.34, 95% CI = 0.14-0.78), but KLK10 polymorphisms were not. Relative expression analysis indicated that an increase in KLK7 and KLK10 mRNA levels was found in cancerous tissues (2(-ΔΔCT) = 25.23 ± 8.85 and 10.89 ± 4.97, respectively). A significantly higher level of KLK7 was expressed in early-stage cancer with the rs10581213(wt/ins + ins/ins) genotypes, but there was no significant difference in the mRNA levels of KLK7 and KLK10 between early- and advanced-stage cancers. CONCLUSIONS: This is the first correlation of OSCC with KLK10 rs3745535G>T polymorphisms. Early-stage OSCC and high KLK7 mRNA levels were correlated with the rs10581213(wt/ins + ins/ins) genotypes. More studies with large sample sizes are needed to verify our findings.

The association of psychosocial factors and smoking with periodontal health in a community population.

BACKGROUND AND OBJECTIVE: The association between psychosocial factors and periodontal disease has been widely reported and might be modified by smoking status. This study investigated the association of periodontal status with psychosocial factors and smoking in a community population. MATERIAL AND METHODS: A structured questionnaire was administered to a total of 1,764 civilian noninstitutional (general population excluding from nursing homes, sanitariums and hospitals) Taiwanese individuals to assess the presence and severity of psychosocial factors [using the 12-item Chinese health questionnaire (CHQ-12)], smoking habits and other related factors. Periodontal status was established using the community periodontal index and by measuring clinical loss of attachment. RESULTS: Psychological factors and smoking were significantly associated with loss of attachment (odds ratio = 1.69, 95% confidence interval = 1.01-2.77, comparing the CHQ-12 score of >or= 6 with the CHQ-12 score of 0-2 and p = 0.032 for linear trend; odds ratio = 2.21, 95% confidence interval = 1.45-3.37, comparing smokers with nonsmokers) but not with community periodontal index. The association was found to be stronger among smokers than among nonsmokers. Smokers with a CHQ-12 score of >or= 6 had a higher odds ratio of loss of attachment (odds ratio = 2.49, 95% confidence interval = 0.91-6.49) than nonsmokers (odds ratio = 1.43, 95% confidence interval = 0.76-2.58). For periodontal health measured using the community periodontal index, married and divorced/widowed subjects tended to have poorer periodontal health (odds ratio = 3.38, 95% confidence interval = 1.26-10.81 and odds ratio = 3.83, 95% confidence interval = 1.21-13.83, respectively) than single subjects among nonsmokers but not among smokers. CONCLUSION: Poor mental health had a stronger association with periodontal disease among smokers than among nonsmokers, especially in accumulative attachment loss. Our findings suggest that mental health and smoking might have a synergistic effect on the risk of developing periodontal disease.

Ethnic differences in the occurrence of oropharyngeal cancer in Taiwan.

OBJECTIVE: The purpose of this study was to examine the oropharyngeal cancer pattern among different ethnic groups in Taiwan. METHODS: The sample population was divided into three ethnic groups: the Fukkien, Hakka, and aboriginal communities. Age-standardized mortality rates (SMRs) and age-standardized incidence rates (SIRs) were estimated among these ethnic groups for the period 1979-1996/1997. RESULTS: Our study found that the higher oropharyngeal cancer mortality and incidence rates in females of aboriginal groups are statistically significant, and higher than reference groups for both genders (SMR=3.76, SIR=2.18). However, in the lower areca quid chewing aboriginal groups, the higher pattern was not seen in females, and the lower pattern was even found in males. The incidence and mortality rate of oropharyngeal cancer in Hakkas was significantly lower than in the reference group. CONCLUSIONS: The pattern of oropharyngeal cancer in Taiwan showed ethnic differences. The differences may be due to variation in exposure to different risk factors; however, in our study, we found that genetic differences might also be considered when explaining the different oropharyngeal cancer patterns among ethnic groups.

Individuals' perception of oral health and its impact on the health-related quality of life.

This research divides oral health status into two parts: clinical disease and patients' subjective perceptions, and then discusses what impact these two items have on the health-related quality of life. A total of 1600 elderly persons, over 65 years old, were sampled as the research target. The overall response rate was 71.62%. SF-36 was used to measure the health-related quality of life, Oral Health Impact Profile (OHIP) was used to measure individuals' subjective perception of their oral health status, and the number of natural teeth was considered as the clinical issue. There are 12 items which affect quality of life in the physical aspect, namely: OHIP score; age; gender; weekly frequency of social activities and interactions; financial status; physician visits per month on a regular basis; hospitalization during the last 6 months; arthritis, rheumatism, cardiovascular disease or diabetes; abnormal IADL patients; and the satisfaction level of current lifestyle. Factors affecting quality of life (i.e. mental aspects), are few, including: OHIP score, gender, education level, weekly frequency of social activities and interactions and satisfaction level of current lifestyle. The results indicate that individuals' subjective perceptions of their oral health status had a greater impact, than did the clinical issue, on their health-related quality of life. The outcomes and findings derived from this research may provide alternative perspectives for professionals, so that they may be more careful concerning individuals' subjective perceptions.

Incidence rates of oral cancer and oral pre-cancerous lesions in a 6-year follow-up study of a Taiwanese aboriginal community.

BACKGROUND: In a previous article, we reported the prevalence rates of oral mucosal lesions in an aboriginal community from an epidemiological survey of oral pre-cancerous lesions. METHODS: Since 1997, the authors have started regular follow-up of the study population originally investigated. Thus, it has been possible to obtain incidence rates for the various oral pre-cancerous lesions and conditions. RESULTS: There were 194 persons without any oral lesion in the 1997 screening. During the clinical follow-up investigation and during the analysis of biopsies from pre-cancerous lesions, we discovered six new lesions (including cancer and pre-cancerous lesions) from five participants. All of the five persons were areca/betel quid chewers, and only one mixed areca/betel quid chewing with cigarette smoking habit. The age-standardized incidence rates for quid lesion, oral submucous fibrosis (OSF) and squamous cell carcinoma (SCC) were 267.0, 374.1 and 146.2 per 100,000 person-years, respectively, for areca/betel quid chewers. CONCLUSIONS: As compared with the rates from India and the general Taiwanese population, the study community encountered a serious problem of oral lesions.

Angiotensin-converting enzyme gene insertion/deletion polymorphism is associated with risk of oral precancerous lesion in betel quid chewers.

To investigate whether angiotensin-converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism is related to the risk of oral precancerous lesions (OPL) in Taiwanese subjects who chew betel quid, a total of 61 betel quid chewers having OPL were compared with 61 asymptomatic betel quid chewers matched for betel quid chewing duration and dosage. The frequency of homozygote for ACE D variant is significantly higher in the case subjects than that of the controls (44.3 vs 24.6%; P = 0.0108). The adjusted odds ratio of the D homozygous for the risk of OPL is 8.10 (95% confidence interval (CI) = 2.04-32.19, P = 0.003). In the allelic base analysis, the D allele is also significantly associated with higher risk of OPL. When grouping the study subjects by smoking status, the association between ACE I/D polymorphism and risk of OPL was only observed in nonsmokers. Our results support the theory that genetic factors may contribute to the susceptibility of OPL and suggest that smoking and genetic factors may be differently involved in the development of OPL.

The effects of chewing areca/betel quid with and without cigarette smoking on oral submucous fibrosis and oral mucosal lesions.

OBJECTIVES: The purpose of this study was to investigate the risk of areca/betel quid chewing with or without cigarette smoking on oral submucous fibrosis (OSF) and other oral mucosal lesions. METHODS: A stratified case-control study was designed. There were in total 102 patients with oral mucosal lesions or OSF (confirmed pathologically) in the case group. OSF (n = 62) and oral mucosal lesions (n = 62) in 102 subjects were separately analyzed for men and women investigating their risks. RESULTS: For OSF, people with both smoking and chewing habits had a statistically significant odds ratio (OR) 8.68 (95% CI = 1.87, 40.23). For the group of people with chewing habit only and without any lifetime cigarette smoking habit, the OR was 4.51 (95% CI = 1.20, 16.94). For other oral mucosal lesions, people with mixed habits and chewing only had also significant risks (OR = 8.37 and 3.95, respectively). For both OSF and other oral lesions, the ORs of mixed habits and chewing only were both higher in women than in men. CONCLUSIONS: The areca/betel quid used in Taiwan does not contain any tobacco product. The only way of areca/betel quid could synergize with any tobacco product is through cigarette smoking. A statistically significant association with oral mucosal lesions and OSF was still found in the group of areca/betel quid chewing only.

Cytotoxic effects of dental resin liquids on primary gingival fibroblasts and periodontal ligament cells in vitro.

Cytotoxic effects of resin liquids of three in situ relining dental polymers, Alike, Kooliner, and Tokuso Rebase, and their major components, methyl methacrylate (MMA), isobutyl methacrylate (IBMA), and 1,6-hexanediol dimethacrylate (1,6-HDMA) were investigated. The concentrations of major monomers in these resin liquids were determined by high-performance liquid chromatography. Cellular viability of human gingival fibroblasts (GF) and periodontal ligament (PDL) cells were evaluated by the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide assay. Moreover, patterns of cell death were analysed using annexin V/propidium iodide staining with flow cytometry. The results indicated that Alike liquid contained 91.3% MMA, Kooliner liquid contained 94.5% IBMA, and Tokuso Rebase liquid contained 65.8% 1,6-HDMA. All materials examined had cytotoxic effects on GF and PDL cells in dose-dependent manners. Tokuso Rebase liquid appeared to be the most cytotoxic among the various resin liquids examined. The effects of Kooliner and Tokuso Rebase liquids may have resulted from IBMA and 1,6-HDMA, respectively. Furthermore, the majority of treated cells died from necrosis; whereas a small portion of cells died from apoptosis. In conclusion, the results demonstrated that these liquid forms of dental polymers and their major monomers cause cytotoxic reactions. The direct relining procedure that cures these materials in situ should be used cautiously.

Radiographic alveolar bone loss in untreated Taiwan Chinese subjects with adult periodontitis measured by the digital scanning radiographic image analysis method.

OBJECTIVES: To determine differences in radiographic alveolar bone loss (RABL) by age group, gender and tooth type in subjects with adult periodontitis using the digital scanning radiographic image analysis (DSRIA) method. METHODS: A total of 4262 teeth were examined for RABL in 178 individuals (96 males and 82 females). Mesial and distal sites were determined by the DSRIA method following scanning of periapical radiographs and using the Microstation 95 image software. RESULTS: Linear assessment of RABL using DSRIA revealed the following results. Mean bone loss of maxillary and mandibular anterior teeth was significantly higher (P<0.001 and P<0.01, respectively) in males compared with females using a two-sample t-test. The highest values of mean RABL occurred most commonly in the maxillary first molars (53.8% and 51.4% for females and males, respectively) and the mandibular central incisors (50.4% and 41.5% for males and females, respectively). Mean RABL was greater in males than in females for both the mandibular and maxillary arches, with the exception of molar sites. CONCLUSIONS: Maxillary first molars and mandibular incisors are the highest risk sites. Moreover, greater loss occurs at the molar site in females compared with males for untreated periodontitis in Taiwan Chinese people.

The precancer risk of betel quid chewing, tobacco use and alcohol consumption in oral leukoplakia and oral submucous fibrosis in southern Taiwan.

In areas where the practise of betel quid chewing is widespread and the chewers also often smoke and drink alcohol, the relation between oral precancerous lesion and condition to the three habits is probably complex. To explore such association and their attributable effect on oral leukoplakia (OL) and oral submucous fibrosis (OSF), a gender-age-matched case-control study was conducted at Kaohsiung, southern Taiwan. This study included 219 patients with newly diagnosed and histologically confirmed OL or OSF, and 876 randomly selected community controls. All information was collected by a structured questionnaire through in-person interviews. A preponderance of younger patients had OSF, while a predominance of older patients had OL. Betel quid chewing was strongly associated with both these oral diseases, the attributable fraction of OL being 73.2% and of OSF 85.4%. While the heterogeneity in risk for areca nut chewing across the two diseases was not apparent, betel quid chewing patients with OSF experienced a higher risk at each exposure level of chewing duration, quantity and cumulative measure than those who had OL. Alcohol intake did not appear to be a risk factor. However, cigarette smoking had a significant contribution to the risk of OL, and modified the effect of chewing based on an additive interaction model. For the two oral premalignant diseases combined, 86.5% was attributable to chewing and smoking. Our results suggested that, although betel quid chewing was a major cause for both OL and OSF, its effect might be difference between the two diseases. Cigarette smoking has a modifying effect in the development of oral leukoplakia.

Evaluation of a self-rating screening test for areca quid abusers in Taiwan.

Areca quid chewing is a popular habit and areca is a well-known ethnopsychopharmalogic agent in southeast Asia. While the chewing habit is legal and also socially acceptable in many places of Taiwan, the public health problem of high oral cancer incidence has remained a priority on the health care list in our local health department. Helping areca quid chewers to reduce or even stop the habit will be paramount in the oral cancer prevention programme. Hence, in order to identify the appropriate strategy for stopping the chewing habit, it is important to distinguish whether an areca quid chewer has reached the level of substance abuse.In accordance with the Diagnostic and Statistical manual of Mental disorder (fourth edition, DSM-IV), we developed a specific self-report questionnaire modified from the famous SCAN system, DSM-IV and ICD-10. The initial screening test for areca quid abusers had 52 questions. Its components included the onset age and frequency, subjective craving and feeling, social problems, physical problems, oral symptoms, psychological and abstinence-related problems, the motivation and capacity to abstain, and demographic data. The answers were divided into 'Yes' or 'No'. One hundred and twenty-five areca quid users (53 men, 72 women) were recruited. The abusers tended to have older age, less education, and higher daily consumption of areca quid. There were no differences on motivation to quit chewing (abstinence) between abusers and non-abusers. There were no statistical differences on tobacco-smoking and alcohol-drinking behavior. Based on the statistical analysis of receiver operation characteristic (ROC) curves, 11 questions were chosen for the Self-report Screening Test for Areca quid Abuser (SSTAA). An areca quid chewer's answers with a score of 4 or more in these 11 questions would be considered an areca quid abuser. The modified process of SSTAA is performed for the evaluation of the native culture-related substance user. At this current stage, the SSTAA developed from the study is the first instrument for screening areca quid abuser.

Functional analyses of natural killer cells in macaques infected with neurovirulent simian immunodeficiency virus.

Clearance of HIV and SIV from the peripheral blood by the cellular immune system lessens the viral burden in infected individuals and may have an impact on virus infection of the CNS and the development of CNS lesions. However, the role of immune responses in preventing or limiting CNS infection has not been clearly defined. We investigated the role of natural killer cells in the outcome of SIV infection of macaques as a model for humans with AIDS and HIV encephalitis. In our study, six pig-tailed macaques were infected with the neurovirulent virus, SIV/17E-Fr, and the immunosuppressive virus, SIV/ DeltaB670, in a model system that causes rapid progression to AIDS and a high frequency of CNS lesions. NK lytic activity in each macaque was monitored longitudinally. In addition, we enumerated NK cells and tested macaque PBMC for the ability to lyse SIV-infected target cells. We found that there was a significant inverse correlation (P=0.02) between the robustness of NK response and the development of CNS lesions. Animals lacking strong NK cell responses developed more severe CNS lesions than those with robust NK responses did. Furthermore, pre-infection levels of NK activity were predictive of CNS lesion severity. The macaque with the most robust pre-infection NK activity developed no CNS lesions. In these infected macaques, NK activity was shown to be directed against SIV-infected cells. We extended these in vivo findings to delineate precisely which cell type was mediating this SIV-directed lysis. We used both macaque and human cells to demonstrate that the population that mediated anti-SIV and anti-HIV cytolytic effects was NK cells. Furthermore, we showed that this anti-SIV and anti-HIV cytolytic effect was directed at the envelope protein and not gag proteins. Thus, NK cells have the capacity to recognize and lyse cells expressing SIV and HIV antigens. These data support a role for NK cells in the modulation of CNS disease.

The effects of arecoline on the release of cytokines using cultured peripheral blood mononuclear cells from patients with oral mucous diseases.

In Taiwan there is a significant correlation between oral precancer diseases and oral cancer associated with the betel quid chewing habit. The carcinogenic components of betel quid are arecoline, arecaidine and safrole. However, it is unknown whether these substances influence the immune functions. This study investigated the effects of betel quid on the immune system using cultured peripheral blood mononuclear cells from patients with oral mucous diseases. In our experiment, mononuclear cells from 10 normal persons, 12 patients with precancer lesions, and 16 patients with squamous cell carcinoma were separated from blood samples and cultured. After stimulation by arecoline, the amounts of IL-2, TNF-alpha, TGF-beta and IFN-gamma secreted by mononuclear cells were measured using the ELISA method. The results showed that IL-2, TNF-alpha, and TGF-beta were significantly lower in mononuclear cells of normal persons as stimulated by arecoline. The TGF-beta amount in cells from oral submucous fibrosis patients with betel quid chewing habit (OSF-B) was lower than normal persons or patients who had long term betel quid chewing habit but were without oral mucosal diseases (N-B), and was also lower than the squamous cell carcinoma with betel quid chewing group (SCC-B). TNF-alpha was significantly lower in the squamous cell carcinoma with long term betel quid chewing group (SCC-B) than in normal persons. TNF-alpha was significantly higher in the squamous cell carcinoma without betel quid chewing group (SCC-N) than in normal persons and SCC-B groups. In addition, IFN-gamma was significantly lower in patients who had long term betel quid chewing but were without oral mucous lesions than the normal person and the OSF group. The results proved that betel quid influences cytokines production by mononuclear cells.

Epidemiological survey of oral submucous fibrosis and leukoplakia in aborigines of Taiwan.

A population-based survey was designed to investigate the prevalence of areca/betel quid chewing, oral submucous fibrosis and leukoplakia in a typical aboriginal community of southern Taiwan. Three hundred and twelve people 20 years of age or older were collected in the study. The prevalence of chewing areca/betel quid was 69.5%, with an average of 17.3 portions a day for an average 24.4 years. More women (78.7%) than men (60.6%) chewed areca/betel quid. The prevalences of oral submucous fibrosis and leukoplakia were 17.6% and 24.4%, respectively. It was found that the odds ratio for chewing areca/betel quid and having at least one of the above oral mucosal lesions was 8.21. Any additional smoking or drinking habits were not significant for having oral mucosal lesions. Although the areca/betel quid in Taiwan does not contain any tobacco, a significant association was still identified between areca/betel quid chewing and oral mucosal lesions.

Internal packing of helical membrane proteins.

Helix packing is important in the folding, stability, and association of membrane proteins. Packing analysis of the helical portions of 7 integral membrane proteins and 37 soluble proteins show that the helices in membrane proteins have higher packing values (0.431) than in soluble proteins (0.405). The highest packing values in integral membrane proteins originate from small hydrophobic (G and A) and small hydroxyl-containing (S and T) amino acids, whereas in soluble proteins large hydrophobic and aromatic residues have the highest packing values. The highest packing values for membrane proteins are found in the transmembrane helix-helix interfaces. Glycine and alanine have the highest occurrence among the buried amino acids in membrane proteins, whereas leucine and alanine are the most common buried residue in soluble proteins. These observations are consistent with a shorter axial separation between helices in membrane proteins. The tight helix packing revealed in this analysis contributes to membrane protein stability and likely compensates for the lack of the hydrophobic effect as a driving force for helix-helix association in membranes.

The consistency and reliability of periodontal bone level measurements using digital scanning radiographic image analysis--a pilot study.

The purpose of the present study was to evaluate the consistency and reliability of alveolar bone height measurement using digital scanning radiographic image analysis (DSRIA). A randomly selected (standardized paralleling technique) periapical radiographs of 20 molar teeth (10 maxillary molars and 10 mandibular molars) from 178 individuals with periodontitis were reporting or referred to the patient population of the dental clinics of the teaching hospital. Radiographic measurements were performed and read by two examiners, with a separation time of three weeks of different tooth groups and as established by double reading of 20 molar teeth in 10 individuals using the DSRIA. The calculating data of radiographic alveolar bone loss (RABL) measured and analyzed by the DSRIA for each molar group were compared based on the inter- and intra-examiners' data. The means and standard deviations were calculated to compare radiographs of the inter- and intra-examiners' groups. The reliability coefficients were computed to assess the consistency and reliability for each molar group. The results revealed that the intra- and inter-examiners' reliability coefficients ranged from 0.986 to 0.995 (p < 0.001, significantly different from 0). An excellent reproducibility was indicated in maxillary molar, mandibular molar and of both arches, respectively. It was concluded that the standard periapical radiograph using the DSRIA has the potential to be a valuable and reliable method in measuring linear alveolar bone defects caused by periodontitis.