Potential Effect for the Smoking on Periodontitis From the Perspective of Arginine Metabolites

Study Overview
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Brief Summary
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Arginine metabolites are amino acids that are associated with vascular tone regulation and the level of inflammation, with critical roles in the synthesis of NO. Our aim was to determine the ADMA, SDMA, L-NMMA, L-arginine, L-homoarginine and IL-6 levels in saliva and serum samples from periodontitis patients and periodontally healthy individuals and to assess the levels of these compounds according to smoking status and compare these levels to those of healthy individuals.

Detailed Description
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The study consisted of four groups: healthy individuals (control (C); n=20), smokers with healthy periodontium (S-C; n=20), nonsmokers with Stage III Grade B generalized periodontitis (P; n=20) and smokers with Stage III Grade C generalized periodontitis (S-P; n=18). Clinical periodontal parameters were measured. The determination of methylated arginine metabolites was carried out by LC-MS/MS.

Official Title
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Potential Effect for the Smoking on Periodontitis From the Perspective of Arginine Metabolites Symmetric Dimethylarginine (SDMA) and Asymmetric Dimethylarginine (ADMA)

Conditions
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Periodontitis, Smoking, Inflammatory Response

Intervention / Treatment
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* Other: Saliva and serum sampling


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: systemically healthy clinical diagnosis of periodontitis clinical diagnosis of periodontal health For smoking group the so kerr were denied as smoking at least 10 cigarettes per day and the duration should more than 10 years. Exclusion Criteria: history of regular use of systemic antibiotics anti-inflammatory, or antioxidant drugs (previous 3 months) nonsurgical periodontal treatment (previous 6 months) surgical periodontal treatment (previous 12 months) presence of<10 teeth current medications affecting gingival health (calcium channel blockers, phenytoin, cyclosporine, and hormone replacement therapy) diabetes diagnosis of rheumatoid arthritis pregnancy lactating excessive alcohol consumption.

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 65 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Prevention
Allocation: Randomized
Intervention Model: Single Group Assignment
Interventional Model Description: Saliva and serum of periodontitis and smoking patients collected for analyzing arginine metabolites.
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Active Comparator: Saliva and serum collection of patients and samples molecules analysis<br>aliva and serum sampling Saliva were collected to analyze the selected markers as unstimulated samples during the early hours of the day. The saliva was centrifuged and then transferred into Eppendorf tubes. Venous puncture was performed after saliva collection and 10 mL of blood samples were collected by qualified staff (MY,EY) from each participant. Saliva and serum were then stored at -80 °C until analysis. | Other: Saliva and serum sampling<br>* Saliva were collected to analyze the selected markers as unstimulated samples during the early hours of the day. The saliva was centrifuged and then transferred into Eppendorf tubes. Venous puncture was performed after saliva collection and 10 mL of blood samples were collected by qualified staff (MFD) from each participant. Saliva and serum were then stored at -80 °C until analysis.<br>|
| Experimental: Salivary and serum arginine metabolites ADMA and SDMA observation<br>IL-6 levels in collected samples were determined by ELISA kits and analyzed according to manufacturers' instructions, with colorimetric assessment performed using a microplate reader at 450 nm with the assay detection range between 7.8 and 500 pg/mL. Concentrations were determined based on the respective assay standard curve. All samples were analyzed in duplicate, and the average was used in subsequent calculations. Determination of methylated arginine metabolites: The ADMA, SDMA, homoArg, arginine and L-NMMA levels in saliva and serum were assessed by a liquid chromatography-mass spectrometry (LC MS/MS)* method, which was a modification of the method of Di Gangi et al. | Other: Saliva and serum sampling<br>* Saliva were collected to analyze the selected markers as unstimulated samples during the early hours of the day. The saliva was centrifuged and then transferred into Eppendorf tubes. Venous puncture was performed after saliva collection and 10 mL of blood samples were collected by qualified staff (MFD) from each participant. Saliva and serum were then stored at -80 °C until analysis.<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Pocket probing depth | Measurement of the depth of a sulcus or periodontal pocket determined by measuring distance from a gingival margin to the base of the sulcus or pocket with a calibrated periodontal probe | 6 months | 
| Clinical attachment level | Clinical attachment level (or loss, CAL) is a more accurate indicator of the periodontal support around a tooth than probing depth alone. CAL is measured from a fixed point on the tooth that does not change, the CEJ. To calculate CAL, two measurements are needed: distance from the gingival margin to the CEJ and probing depth. In recession: probing depth (+) gingival margin to the CEJ (add). In tissue overgrowth: probing depth (-) gingival margin to the CEJ (subtract) | 9 months | 
| Bleeding on probing | referring to bleeding that is induced by gentle manipulation of the tissue at the depth of the gingival sulcus, or interface between the gingiva and a tooth | 9 months | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Saliva and serum samples processing | Phenomenex (Torrance, CA, USA) 75 x 4.6 mm x 4 µm polar-RP column, Dionex HPLC and Access MAX LC-MSMS (Thermo Scientific, USA) device were used; in the gradient phase mobile phase A (40 mM ammoniumformate, 3% formic acid) and mobile phase B (acetonitrile) were utilized. A flow chart with increasing in steps of 2-8 minutes for 10% B phase, 30% B phase for 8-11 minutes, and finally 10% B phase for 11-20 minutes was applied at a flow rate of 0.3 ml/min. MRM and CE values were as follows: Arginine 231.3-70.1 CE: 15 V; Homoarginine 245.2-84.2 CE: 24 V; L-NMMA 245.3-70.2 CE:15V; ADMA 259.3-214.0; CE:25V; SDMA 259.3-228.0 CE:15V; ADMA-D7 266.610-221.000 CE:15V; capillary temperature 210 °C; Vaporizer temperature 350 °C; Sheath gas pressure: 30 Arb.; Aux Gas pressure:10 Arb.; Ion Sweep Gas Pressure: 0; Spray Voltage: 3000 V; Polarity: (+). | 2 months | 
| serum and saliva analyses for arginine metabolites | The ADMA, SDMA, homoArg, arginine and L-NMMA levels in saliva and serum were assessed by a liquid chromatography-mass spectrometry (LC MS/MS)* method, which was a modification of the method of Di Gangi et al. | 2 months | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Periodontitis, Arginine metabolites, Smoking