SUMMARY AND FURTHER THOUGHTS

15 May

The factors involved in the pathogenesis of acne are depicted in Figure 11 (125), and although the  remission of acne  remains a mystery (125,126), several  technologies have  been  developed for the  topical  control  of acne  identified through single  in vitro assays  or a combination of different models.

Benzoyl peroxide has been reported to have multiple effects including bacter- iostatic activity, mild comedolytic activity,  enhancement of local blood flow, and the release  of free  radical   oxygen.  Benzoyl  peroxide is  an  organic   peroxide that  is bactericidal  against  P.  acnes  along   with   additional  anti-inflammatory  activity (127).  There  is  a  wide   range   of  antimicrobial activity   against multiple species and  resistance does  not  occur.  Hegemann et al. (127) investigated the  release  of reactive oxygen   species   regulated by  protein kinase   C  (PKC)  and   calmodulin from   human  neutrophils;  although  benzoyl  peroxide  inhibited  PKC  activity (IC50   of 1.35 mM),  there  was  no  activity  against calmodulin, suggesting that  the

FIGURE 11    Synopsis of factors  involved  in the  pathogenesis of acne. Factors involved  in the pathogenesis of acne are  increased sebum production, ductal  cornification,  bacterial colonization, and  inflammation.  Abbreviations: HLA-DR, human  leukocyte antigen-D  related;  ICAM, intercellular adhesion molecule; IL, interleukin; TNF, tumor necrosis factor. Source: From Ref. 125.

anti-inflammatory  activity   of  benzoyl peroxide  was  not  mediated by  PKC  or calmodulin.

Babich et al. (128) used  a human keratinocyte cell line to evaluate the cytotox- icity of benzoyl peroxide. As determined by uptake of Neutral red, irreversible cell death was evident after a one-hour exposure to 0.15 mM and greater concentrations of  benzoyl peroxide, whereas reductions in  proliferation were  seen  at  concen- trations from  0.02 to  0.08 mM.  Stress  and  damage as  evident by  vacuolization and  LDH  release  were  seen  after  four  hours at 0.05 mM  levels.  As a control,  the IC50  of benzoic  acid was  29.5 mM.

Similar  degrees of  cytotoxicity were  found with   other  organic   peroxides

suggesting that  the  cytotoxicity of benzoyl peroxide may  be related to the  gener- ation  of reactive oxidative free radicals. King et al. (129) found that  benzoyl per- oxide,  as a free radical generating compound, acts as a tumor promoter in mouse skin  by  causing DNA  strand breaks  and  base  modifications in  cultured murine keratinocytes. Matsui et al. (130) also found that  benzoyl peroxide inhibited both human and  murine PKC in a cell-free system, although there  was no translocation evident from the cytosol to the membrane when tested in cultured human keratino- cytes.  Other  reports investigating the  mechanism of action  of benzoyl peroxide include Valacchi  et al. (131) who  examined the  depletion of vitamin E and  IL-1a gene   expression  in  HaCaT   keratinocytes,  Burkhart  et  al.  (132)  who   reported increases in  free  radical activity  in  combination with  antibiotics, and  Lawrence et  al. (133) in  that  benzoyl peroxide interferes with  metabolic cooperation with cultured keratinocytes.

Another compound that has been utilized for the topical  control  of acne is sal- icylic acid. Salicylic acid reduces inflammation by inhibiting the synthesis of pros- taglandins that  are  generated in inflamed tissues.  Salicylic  acid  also  inhibits the conversion of arachidonic acid  to PGE2, which  is catalyzed by the enzyme cyclo- oxygenase. Salicylic acid also inhibits comedogenesis by promoting desquamation of the  follicular  epithelium. Additional agents that  have  been  used  to treat  acne include sulfur for the treatment of inflammatory acne lesions  through keratolysis activity  and azelaic acid, a dicarboxylic acid that has both antibacterial and antiker- atinizing activity  (134,135).

In addition to the models and  assays  described in this review,  other  in vitro models have  been  developed around  the  hair  follicle.  Although these  models were  not developed to look at acne per se, they  may  be of interest for specific cell types. Blume et al. (136) reported on the monolayer culture of keratinocytes isolated from  terminal hair  follicles  that  express glycoproteins during proliferation (gp38) and  differentiation (gp80). These glycoproteins are absent in normal interfollicular keratinocytes. Limat  and  Noser  (137) also  reported on  the  serial  cultivation of keratinocytes from  the outer  root sheath of hair  follicles.

Lenoir-Viale et al. (138) documented the effects of retinoic  acid on an in vitro model  reconstructed from  the  outer  root  sheath of human hair  follicles,  whereas Hoeller  et al. (139) divulged an improved method on the  construction of in vitro skin  equivalents from  human hair  follicles  and  fibroblasts. Additionally, Michel et al. (140) has  developed a tissue-engineered human skin  equivalent containing hair  follicles.

The in situ  organ  culture of the entire  pilosebaceous appendage has  further been   developed  by  Kealey’s   laboratory  to  monitor  hair   growth.  This  model appears to  be  the  most  complex,  as  it  contains the  entire  follicle  including the sebaceous gland  (141 – 143).

Katz  and  Taichman (144) reported on  the  development of a two-chamber culture model  in which  proteins secreted by keratinocytes could  be isolated and characterized.  In  this   model,  a  fully  differentiated  keratinocyte  epithelium  is grown on plastic  inserts apart from fibroblasts in the lower  chamber.

In conclusion, multiple in vitro  models are available for the identification of technologies to alleviate or prevent the  formation of acne  lesions.  By using  these models, it is possible to determine the mechanism of action  of lead  candidates.

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