Autophagy, One of the Main Steps in Periodontitis Pathogenesis and Evolution.

Periodontitis represents a complex inflammatory disease that compromises the integrity of the tooth-supporting tissue through the interaction of specific periodontal pathogens and the host's immune system. Experimental data help to outline the idea that the molecular way towards periodontitis initiation and progression presents four key steps: bacterial infection, inflammation, oxidative stress, and autophagy. The aim of this review is to outline the autophagy involvement in the pathogenesis and evolution of periodontitis from at least three points of view: periodontal pathogen invasion control, innate immune signaling pathways regulation and apoptosis inhibition in periodontal cells. The exact roles played by reactive oxygen species (ROS) inside the molecular mechanisms for autophagy initiation in periodontitis still require further investigation. However, clarifying the role and the mechanism of redox regulation of autophagy in the periodontitis context may be particularly beneficial for the elaboration of new therapeutic strategies.

Salivary biomarkers: relationship between oxidative stress and alveolar bone loss in chronic periodontitis.

OBJECTIVES: Oxidative stress is implicated in the pathogenesis of many systemic and oral diseases such as periodontal disease. The main aim of this study is to explore a possible association between salivary markers of OS and alveolar bone loss. MATERIALS AND METHODS: The study included 20 patients with chronic periodontitis and 20 controls. Salivary OS biomarkers 8-hidroxy-desoxguanosine (8-HOdG), malondialdehyde (MDA), uric acid, total antioxidant capacity (TAC) and glutathione peroxidase (GPx) were evaluated. Bone loss markers such as C-terminal telopeptide of type I collagen (CTX I), matrix metalloproteinases-8 (MMP-8), osteocalcin and 25-hydroxy vitamin D3 (25- OH D) were detected in this study. The methods included general biochemical tests and ELISA. RESULTS: Salivary 8-OHdG, MDA levels were significantly higher in the chronic periodontitis group compared with controls (p < 0.05). Salivary activities for uric acid, TAC and GPx were significantly decreased in patients with chronic periodontitis vs controls (p < 0.05). Salivary levels for CTX I, MMP-8, 25-OH D and Osteocalcin were significantly higher in the chronic periodontitis group compared to the controls (p < 0.05). A significant positive correlation was observed between salivary levels of MDA and CTX I. Significant negative correlations between uric acid and CTX I and between MMP-8 and uric acid have been found. Significant positive correlations were observed between CTX I, MMP-8, 25-OH D, osteocalcin and clinical parameters of periodontal disease. CONCLUSIONS: Important oxidative stress associated with alveolar bone loss biomarkers can be detected in saliva of patients with periodontal disease.

Effect of diet and omega-3 fatty acids in NAFLD.

Nonalcoholic fatty liver disease (NAFLD) ranges from simple hepatic steatosis to steatohepatitis (NASH) and cirrhosis. The aim of this study is to test beneficial effects of omega-3 fatty acids (DHA 130 mg, EPA 25 mg) treatment in NAFLD, in a mouse model of non-alcoholic fatty liver disease. As pretreatment, 50 mice were fed for one month with a high-fat diet to induce NAFLD. Then, the mice were divided in different groups according to diet (normo- or hypercaloric), with and without treatment with omega-3 fatty acids, for another month, forming the post-treatment group. The liver and blood samples were collected for biochemical and histopathological analysis. Biochemical parameters including: glycemia, total cholesterol, triglycerides, uric acid, albumin, total plasma antioxidant capacity (TEAC) was measured in serum. Glutathione (GSH), total thiols and malonyldialdehyde (MDA) were determined in mouse liver homogenates. Mice from post-treatment group, on hypercaloric diet with or without omega-3 fatty acids treatment, had medium hepatopathy (granular and vacuolar degeneration of the hepatocytes) and hypertriglyceridemia. Omega-3 fatty acid treatment lowered the rise of triglycerides (p<0.03), glycemia (p<0.01) and cholesterol (p<0.02) in serum and MDA level of the liver (p<0.05). Mice from post-treatment group, on normocaloric diet with or without omega-3 fatty acid had different histopathological and biochemical results. Those with normocaloric/normolipidic diet and omega-3 fatty acids treatment had reversed liver histopathological results from NASH to normal aspect and had the best metabolic parameters results. In conclusion, omega-3 fatty acids treatment associated with a normocaloric/normolipidic diet has hepatoprotective action in nonalcoholic fatty liver disease.

Diet and paraoxonase 1 enzymatic activity in diabetic foot patients from Romania and Belgium: favorable association of high flavonoid dietary intake with arylesterase activity.

BACKGROUND/AIMS: The antiatherosclerotic enzyme paraoxonase (PON1) is affected by disease and lifestyle. We investigated the impact of diet in diabetic foot patients from 2 European countries. METHODS: Dietary intake and serum PON1 activity, using as substrate paraoxon (paraoxonase) or phenylacetate (arylesterase), were assessed in patients from Bucharest (n = 40) and Antwerp (n = 30) and in 34 healthy controls. RESULTS: The diabetic patients had lower paraoxonase and arylesterase activities than the controls. Arylesterase was lowest in the Bucharest patients, 116 +/- 42 U/ml, versus 141 +/- 43 and 184 +/- 49 U/ml in the Antwerp patients and controls, respectively (p < 0.0005). The Bucharest patients had worse glycemic control, higher blood pressure, lower HDL cholesterol and a diet richer in cholesterol and poorer in monounsaturated fats and fish. In contrast, their median intake of vitamins E and C, folic acid and flavonoids was higher, 82 mg (range: 4-259 mg), versus 28 mg (range: 5-169 mg) aglycone units in Antwerp (p = 0.005). Flavonoid intake predicted arylesterase independently of HDL cholesterol, region and sex (beta = 0.27; p = 0.03), and patients with high intake achieved normal levels of arylesterase (30.1 +/- 10.0 U/micromol in the highest versus 21.0 +/- 8.2 U/micromol total cholesterol in the lowest tertile; p = 0.02). CONCLUSION: A flavonoid-rich diet is positively associated with PON1 arylesterase activity in diabetic foot patients.

Hepatocyte steatosis in patients infected with genotype 1 hepatitis C virus.

BACKGROUND: Recent findings suggest a higher prevalence of hepatic steatosis in patients with chronic hepatitis C, estimated at 50%. Both host and viral factors contribute to the development of steatosis in chronic hepatitis C. Steatosis is an initial stage, which promotes hepatic fibrosis through oxidative stress. AIM: To assess the pathogenic mechanism of genotype 1 hepatitis C virus in steatosis and to evaluate the correlation between the degree of steatosis and the level of oxidative stress. PATIENTS AND METHODS: The study was carried out on 50 patients (29 males, 21 females) with genotype 1 HCV and liver biopsy proven chronic hepatitis C. Patients with other etiology of chronic liver disease were excluded. We statistically correlated the degree of steatosis with clinical (age, sex, waist circumferences) and biological parameters (alaninaminotransferase, gammaglutamyltranspeptidase - GGT, insulin, ferritin, serum viral load, oxidative stress). Insulin resistance (IR) was determined by the homeostasis model assessment (HOMA) method. The oxidative stress was estimated by serum malondialdehyde (MDA) and glutathione (GSH). RESULTS: 27 patients presented steatosis (57%): 14 out of 29 men (48%) and 14 out of 21 women (66%); in two thirds of them, steatosis was moderate. Univariate analysis identified five parameters that significantly influenced steatosis: age >45 years, sex - female, IR (HOMA>2.5), BMI, central adiposity (as reflected by waist circumferences and high GGT-values). Multivariate analysis identified four significant parameters: sex - female, insulin resistance (HOMA>2.5), BMI>30 kg/m(2) and GGT>2N. No relationship was found between steatosis and viral replication. The study demonstrated a significant correlation between steatosis and IR on the one hand and between steatosis and liver fibrosis on the other hand (p<0.05). Liver fibrosis was significant correlated with the increase levels of free radicals (MDA>250 nmol/dL). CONCLUSIONS: The pathogenic mechanism of genotype 1 HCV in steatosis is independent from viral replication and it may be linked to virus induced metabolic abnormalities such as IR. More women (66%) than men (48%) developed steatosis. Increased levels of free radicals, correlated with moderate and severe steatosis suggest the intervention of oxidative stress in determining the hepatic lesions associated with steatosis.

Role of oxidative stress in the pathogenesis of chronic hepatitis C (CHC).

UNLABELLED: Oxidative stress is an important pathophysiological mechanism in chronic hepatitis with C-virus infection (CHC). Steatosis is frequently observed in CHC and seems to have a significant impact on the natural history of the disease with respect to development of fibrosis. The aim of this study was to investigate the relationship between systemic parameters of oxidative stress, insulin resistance, steatosis degree, and fibrosis in CHC. Fifty patients with chronic hepatitis C (29 men and 21 women with the average age 45 years), with or without steatosis, were tested for: oxidative stress and antioxidant status by measuring serum malondialdehyde (MDA), total blood non-proteic thiols concentration (GSH), gamma glutamyl transpeptidase (GGT) activity, lipid parameters, and liver function tests. Our results show that the prevalence of insulin resistance (IR) in chronic hepatitis with C-virus genotype 1 was 32% and the association with hepatic steatosis was in a proportion of 48%, IR is mediated by both metabolic factors as well as viral factors. Hepatic steatosis was associated with an increase of MDA, correlated with its severity, and secondary with a decrease of GSH. The activity of serum GGT was net superior, in patients with steatosis, proportional with its degree. CONCLUSIONS: In patients infected by HCV genotype 1, oxidative stress and insulin resistance contribute to steatosis, which in turn exacerbates both insulin resistance and oxidative stress and accelerates the progression of fibrosis. The induction of GGT is an adaptive response against oxidative damage elicited by lipid peroxidation and it may be critical in the progression of the disease.

Saliva--a diagnostic window to the body, both in health and in disease.

Saliva, the most available and non-invasive biofluid of the human body, permanently "bathes" the oral cavity and is trying to cope with an ever-changing milieu. The oral cavity, a very complex and unique milieu due to its dual function, is the only place in the body where the mineralized tissue is exposed to the external environment in which there are complex interactions between various surfaces: host soft and hard tissues, food, air, and microorganisms. Saliva includes a large number of inorganic and organic compounds, which act as a "mirror of the body's health." In addition to its other functions, saliva could constitute the first line of defense against oxidative stress. Due to its composition and functions, saliva could have a significant role in controlling and/or modulating oxidative damages in the oral cavity. As a diagnostic fluid, saliva offers distinctive advantages over serum. Furthermore, saliva may provide a cost-effective approach for the screening of large populations. Gland-specific saliva can be used for diagnosis of pathology specific to one of the major salivary glands. Whole saliva, however, is most frequently used for diagnosis of systemic diseases. As we enter the era of genomic medicine, sialochemistry will play an increasingly important role in the early detection, the monitoring and progression of the systemic and oral diseases. We reviewed the current data within literature and of our research concerning clinical potential of the saliva.

Oxygen, a paradoxical element?

Oxygen is an essential element for life on earth. No life may exist without oxygen. But in the last forty years, conclusive evidence demonstrated the double-edge sword of this element. In certain conditions, oxygen may produce reactive species, even free radicals. More, the production of reactive oxygen species (ROS) takes place everywhere: in air, nature or inside human bodies. The paradox of oxygen atom is entirely due to its peculiar electronic structure. But life began on earth, only when nature found efficient weapons against ROS, these antioxidants, which all creatures are extensibly endowed with. The consequences of oxygen activation in human bodies are only partly known, in spite of extensive scientific research on theoretical, experimental and clinical domains.

Genetic testing for multiple endocrine neoplasia type 2.

Multiple endocrine neoplasia type 2 (MEN 2) represents a complex autosomal dominant inherited syndrome characterized by occurrence of distinct proliferative disorders of endocrine tissues. The identification of RET proto-oncogene mutations in MEN 2 and FMCT has provided a precise method for identifying gene carriers. 30 subjects (9 males, 21 females, age range 11-63 years) with multiple endocrine neoplasia type 2 have been investigated from 1998 till 2006. 20 patients were considered as index cases and 10 patients were identified after a screening programme for MEN 2. Tumoral associations permitted the MEN 2A diagnosis in 21 cases, MEN 2A with cutaneous lichen amyloidosis in 6 cases and FMCT in 3 cases. We selected 22 patients from 14 families to investigate mutations in the RET proto-oncogene. In 7 subjects no mutations could be detected in the exons 10 and 11 of the RET proto-oncogene. Heterozygous missense mutations in exon 11 were found in 15 subjects consisting of three different mutations in codon 634 (TGC --> TGG, TGC --> GGC, TGC --> CGC). We conclude that our 15 patients have the most frequent mutations described in MEN 2A families. Because the testing for exons 10 and 11 is negative for other 7 patients, the remaining 13, 14, 15 and 16 exons should be sequenced in these cases.

Cigarette smoke effect on total salivary antioxidant capacity, salivary glutathione peroxidase and gamma-glutamyltransferase activity.

Saliva is the first biological fluid that inhaled cigarette smoke (CS) encounters. CS contains several carcinogens known to initiate and promote tumourigenesis and metastasis. One of the aims of this study was to establish if glutathione peroxidase and gamma-glutamyltranspherase (GGT) could be used as possible markers for evaluating the oral oxidative stress caused by smoking. The effect of CS on free radical generation was investigated using two methods. Using different assays, different antioxidants present in saliva may be evidenced due to the different principles on which they are based. Our results indicate that exposure to CS caused a statistically significant decrease of both salivary glutathione peroxidase (p < 0.01) and salivary GGT (p < 0.01). We also found that exposure to CS caused a statistically significant decrease of salivary total antioxidant status (p < 0.01). Such decreases may have a consistent role in the mechanisms by which the toxic effects of CS initiate oral inflammatory diseases, promote precancerous transformations, and destroy the oral cavity homeostasis. Therefore the evaluation of total antioxidant capacity of saliva is important but it must be done together with the evaluation of salivary specific markers of oxidative stress, such as uric acid, albumin and possibly, GGT.

Effect of gas phase and particulate phase of cigarette smoke on salivary antioxidants. What can be the role of vitamin C and pyridoxine?

The effect of smoking is in our days a serious global public health problem of major concern. Incidence of oral squamous cell carcinoma (SCC) in cigarette smokers is four to seven times higher than in nonsmokers. There is a constant and direct attack of various cigarette smoke constituents on the oral epithelial cells, which gradually accumulate and cause malignant transformation. Saliva is the first biological fluid that encounters inhaled cigarette smoke (CS). We have studied the influence of CS on salivary antioxidant capacity, uric acid, amylase and LDH (lactate dehydrogenase). In our study both, gas and particulate phase of CS were tested separately, and possible antioxidant effect of pyridoxine on salivary components was examined. Our results indicate that exposure to both, gas and particulate phase of CS caused a statistically significant decrease in salivary uric acid, LDH and amylase activity. We have also studied the effect of vitamin C (10 mg/dl) and vitamin B6 (1 mM) during incubation of saliva in the presence of CS. The addition of vitamin C had a significant (p < 0.05) protective effect on salivary uric acid level (0.25 +/- 0.12 for saliva incubated with gas phase of CS vs. 0.65 +/- 0.12 for saliva incubated with gas phase of CS in the presence of vitamin C). Vitamin C was not able to maintain/restore the original uric acid level. In the presence of the gas phase, pyridoxine had no protective effect, neither on salivary uric acid level nor on the FRAP activity of saliva. The purpose of our study was to discover a connection between the level of antioxidants in saliva in the presence of the two components of CS. Our results show that salivary antioxidant system is significantly and distinctly affected by both gas and particulate phase of CS and suggest that an adequate intake of antioxidants may help smokers to avoid CS-induced oxidative damage and to prevent degenerative diseases.

Diabetic foot patients with and without retinopathy and plasma oxidative stress.

OBJECTIVE: The first aim of this study was to evaluate some plasma oxidative stress markers in diabetic patients (type2 diabetes mellitus) with peripheral arteriopathy. Secondly, these patients were divided into two groups considering the presence or absence of retinopathy. The differences in the levels of the oxidative stress parameters could be important to select patients prone to develop multiple vascular complications, including retinopathy. PATIENTS AND METHODS: Forty hospitalized type II diabetic patients, aged between 40 and 70, with stage III or IV foot ulceration according to the Wagner classification, had an ophthalmologic evaluation for retinopathy. Among them, 23 were with retinopathy with different grades of severity and 17 were without retinopathy. Twenty healthy subjects served as a control group. Fasting plasma glucose, glycosylated hemoglobin, serum fructosamine, total serum proteins, serum uric acid and plasma ceruloplasmin levels were determined and compared. RESULTS: Plasma levels for fasting glucose, glycosylated hemoglobin, ceruloplasmin and serum concentrations for fructosamine and uric acid were significantly increased in diabetic foot patients vs. control subjects. Comparing the two groups of patients, with and without retinopathy, the concentrations of ceruloplasmin and uric acid were significantly increased in diabetic patients with retinal disease. In diabetic patients with retinopathy, positive correlations were calculated between glycated hemoglobin and fructosamine concentrations and between glycated hemoglobin and ceruloplasmin. CONCLUSION: Diabetic foot patients with retinopathy have increased plasma levels of uric acid and ceruloplasmin. These plasma compounds could be important in the pathogenesis of retinal disease. Two aspects should be considered when these high values are analysed. First, these antioxidant compounds may become prooxidant in diabetic vascular environment. Secondly, it is not known whether these modified plasma oxidative stress parameters are cause or effect in diabetic complication development.

Could constitute saliva the first line of defence against oxidative stress?

The oral cavity, a very complex and unique milieu due to its dual function, respiratory and digestive, is the only place in the body where the mineralized tissue is exposed to the external environment. In this environment there are complex interactions between various surfaces: host soft and hard tissues, food, air and microorganisms. Saliva is in the middle of this environment and tries to cope with an ever-changing milieu. It includes a large number of inorganic and organic compounds, which act as a "mirror of the body's health". In addition to its other functions, saliva could constitute the first line of defence against oxidative stress. The body contains a number of protective antioxidant mechanisms. Saliva is also rich in antioxidants. We review the current available data and from our laboratory concerning antioxidant capacity of saliva. Substantial data are available in literature on the role of reactive oxygen species and antioxidants in diseases, but few information is available on oral pathology. Due to its composition and functions, saliva could have a significant role in controlling and/or modulating oxidative damages in the oral cavity.

Free radical activity in obese patients with type 2 diabetes mellitus.

UNLABELLED: Oxidative stress plays critical roles in the pathogenesis of various diseases. The aim of the present study was to investigate the relationship between oxidative stress, measured by plasma dicarbonyls and plasma antioxidant defence, measured by reduced glutathione (G-SH) in the whole blood, protein thiols in the plasma and erythrocyte superoxide dismutase activity in obese and obese with type-2 diabetes mellitus subjects, clinically free of complications. Twenty obese patients with type-2 diabetes mellitus and twenty nondiabetic obese patients were examined and compared with twenty healthy controls (matched for age and sex against the obese patients with or without diabetes). RESULTS: Obese patients and obese diabetic patients had lower blood glutathione than control subjects (p<0.02 and respectively p<0.04) and higher plasma MDA, an end product of lipid peroxidation (p<0.004 and respectively p<0.01). There was a significant difference between plasma MDA from obese and obese diabetic subjects (p<0.05). Plasma thiols (expressed as micromol/mg protein) did not differ between the three groups. Plasma dicarbonyls concentrations were significantly increased in obese (p<0.043) and obese diabetic patients (p<0.047) and SOD activity (U/g Hb) was significantly decreased in obese (p<0.0 ) and obese diabetic patients (p<0.05) compared to controls. Analysing the results of our study, which show that most of the markers of oxidative stress are modified in the same way in obesity and obesity with diabetes mellitus type 2, we suppose that obesity leads to oxidative stress which can contribute to obesity-associated diseases such as diabetes mellitus.

A comparative oxidative stress study--obesity with and without diabetes mellitus.

Oxidative stress and inflammation are involved in the initiation and progression of obesity and diabetes mellitus. The aim of our study was to find out some markers of oxidative stress in twenty obese patients with type 2 diabetes mellitus (group D) and twenty age-matched obese subjects (group O) and compare the results with the control values from twenty matched healthiy subjects (group H). Spectrophotometric methods were used. For the following plasma parameters: ceruloplasmin, d-ROM (determinable Reactive Oxygen Metabolites), alpha-dicarbonyls, the values were modified in the same way for the groups of patients versus healthy subjects. The patients had higher alpha-dicarbonyls levels than the controls (for D versus H, p<0.047 and for O versus H, p<0.043). There were not significant differences for plasma ceruloplasmin and d-ROM levels. Comparing group O versus D, all the above parameters had very close values. The antioxidant capacity (AC) was higher in group O versus group H (p<0.001) and higher in group O versus D (p<0.02). The high AC for obese patients may be due to hyperuricemia. A negative correlation between AC and d-ROM concentrations and a positive correlation between ceruloplasmin and AC levels was observed for group D. Our data underline that in type 2 diabetes mellitus and obesity, the plasma markers of oxidative stress are modified in the same way. Oxidative stress may be a "connector" between these two diseases. Probably body fat reduction (for obese individuals) diminishes oxidant formation and, in its turn, the incidence of obesity related diseases, such as diabetes mellitus.

The effect of fasting on the parameters of the antioxidant defence system in the blood of vegetarian human subjects.

OBJECTIVES: Although the evidence is strong that fasting has anti-tumor, anti-inflammatory and anti-ageing actions, the mechanisms responsible for these phenomena are still unclear. An ameliorated antioxidative defence with fasting may be the cause of such effects. The objective of the present work was to determine the influence of fasting on antioxidant systems in healthy young vegetarian humans. DESIGN AND METHODS: We measured Trolox Equivalents Antioxidant Capacity (TEAC) of plasma, erythrocytes superoxide dismutase (SOD) activity, blood glutathione peroxidase (GPx) activity, level of total blood non-proteic thiols (TBNT), plasma ceruloplasmin activity, plasma level of NO metabolites (the sum of nitrites and nitrates, NOx), in 18 healthy young humans (age 20-27 years) after 12h (overnight fasting) and 80h of fasting. RESULTS: Trolox Equivalents Antioxidant Capacity of plasma, the level of total blood nonproteic thiols, plasma ceruloplasmin activity and plasma concentration of nitrites and nitrates were significantly increased after 80h of fasting. Superoxide dismutase activity and glutathione peroxidase activity were lower after 80h of fasting. CONCLUSIONS: Our results suggest that fasting induces the "reorganisation" of antioxidative defence lines: fasting increases especially plasma protective systems (total antioxidant capacity of plasma, plasma ceruloplasmin activity) and decreases an erythrocytes antioxidant enyzme (superoxide dismutase) and blood glutathione peroxidase.