טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentKlein Ifat
SubjectCigarette Smoke Damage to Saliva Defense Systems and the
Capacity of Antioxidants to Curtail Some of these
Damages
DepartmentDepartment of Medicine
Supervisor Professor Abraham Reznick


Abstract

Background: Saliva is the first body fluid to encounter cigarette smoke (CS). Saliva is known for its highly protective functions against deleterious agents such as microorganisms, toxins, and various oxidants. CS contains over 4000 different chemicals, 400 of which are proven carcinogens. CS also contains oxidants such as free radicals and volatile aldehydes, which are probably the major causes of damage to biomolecules exposed to CS.  Among the many of functions of saliva, the most  relevant to this work are those involved in antibacterial and antioxidant defenses. The working hypothesis of this thesis is that exposing saliva to CS will cause severe damage to these defense systems, which may lead to the development of oral pathologies and malignancies known to occur more frequently in smokers than in nonsmokers.  The aims of the present study were: 1) To characterize the antioxidant defense system of human saliva. 2) To study the general effects of CS on salivary proteins and enzyme activities. 3) To characterize the oral lactate dehydrogenase (LDH) and to study the effect of CS on its isoenzyme activities and mechanism of inactivation. 4) To study the effects of exposure to CS on the activity of oral peroxidase (OPO) activities and the mechanisms of his inactivation. Results:  This study showed that exposure of saliva in vitro to CS caused a marked reduction of activities of several salivary enzymes as well as accumulation of protein carbonyls in salivary proteins.  Thiol antioxidants such as glutathione (GSH) and N‑acetylcysteine  (NAC) provided protection to amylase and LDH from CS exposure, indicating that ‑SH groups are involved in the mechanisms of inactivation.  The profile of LDH isoenzymes in saliva was completely different from that of plasma, and the major source of the enzyme was found to be the oral epithelial cells.  CS  exposure resulted in a  marked destruction of LDH isoenzymes, whereas it did not affect plasma LDH enzymatic profiles. CS also caused significant inactivation (about 80%) of OPO, which in vivo was reversible in half an hour.  In vitro studies revealed that none of the antioxidants  used, such as GSH, ascorbic acid, Desferal, etc., could provide any protection, and neither an oxidative system (Fe3+, ASC) nor aldehydes present in CS, could affect OPO activity. Subsequent studies showed that the main factor in CS that  inactivates the OPO was hydrocyanic acid (HCN), present in microgram quantities in CS. Addition of  hydroxocobalamine, a known CN chelator, significantly protected the CS-associated inactivation of OPO. Conclusions: The effects of CS on saliva are multifactorial, with various agents in CS affecting salivary components via different mechanisms. The significance of CS associated loss of peroxidase ctivity, may be a cause of CS-induced oral pathologies such as oral inflammations and malignancies.