Bhatti, Manpreet; (2000) A study of the photolysis of Porphyromonas gingivalis. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Porphyromonas gingivalis has been implicated in the development of periodontal diseases and the current treatments used are not entirely successful in eradicating periodontopathogens from the pocket. Photodynamic therapy has been successfully used in the treatment of a number of cancers and has been used in vitro to kill a variety of bacteria. Lethal photosensitisation of Por. gingivalis using toluidine blue O (TBO) with Helium/Neon (HeNe) laser light was compared to lethal photosensitisation using aluminium disulphonated phthalocyanine (AIPCS2) with gallium aluminium arsenide (GaAs) laser light. It was found that there were substantial reductions in viable counts using TBO/HeNe laser light and when using AIPcS2/GaAs laser light. Substantial kills were obtained when lethal photosensitisation was carried out under conditions most likely to be encountered in the periodontal pocket (presence of serum, increasing pH values, different Por. gingivalis strains and biofilm-grown bacteria). The involvement of singlet oxygen and hydroxyl radicals was investigated by carrying out lethal photosensitisation in the presence of the singlet oxygen enhancer, deuterium oxide. It was found that singlet oxygen and possibly hydroxyl radicals are involved in the killing of Por. gingivalis. Distribution studies using 3H-TBO showed that most of the 3H-TBO was bound to the outer membrane and SDS-PAGE analysis showed that there were alterations to the outer and plasma membrane proteins from cells sensitised with TBO and exposed to laser light. It was also found that there was DNA degradation as a result of lethal photosensitisation. The final part of the study involved conjugating antibody against surface components of Por. gingivalis with TBO and specifically to target Por gingivalis to light-induced killing when in the presence of commensal bacteria or host tissue. When sensitised with the antibody/TBO conjugate and exposed to laser light, there were no significant reductions in viable counts of Streptococcus sanguis or human gingival fibroblasts, whereas all of the Por. gingivalis present were killed. In conclusion, Por. gingivalis can be killed effectively (100%) when sensitised with 82 μM TBO at an energy dose of 4.4 J. Type II and possibly type I mechanisms are involved in the killing of Por gingivalis and the outer and plasma membrane proteins and DNA are adversely affected by lethal photosensitisation. Damage to oral commensal organisms and oral host tissue can be avoided by using laser light in combination with TBO conjugated to antibody against Por gingivalis surface components.

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