1. Decreased Extracellular-Signal-Regulated Kinase and Increased Stress-Activated MAP Kinase Activities in Aged Human Skin In Vivo 
Jin Ho Chung, Sewon Kang, James Varani, Jiayuh Lin, Gary J. Fisher, John J. Voorhees 
Journal of Investigative Dermatology 
Volume 115, Issue 2, Pages (August 2000)
DOI: /j x
Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
ERK signaling pathway is decreased in old human skin in vivo. (a) Total (▪) and phosphorylated ERK (□) protein were measured in soluble skin extracts from young (mean age 23.8 y, N=12) and old (mean age 84.3 y, N=12) persons by Western blot. Insets show representative Western blots from two young (1 and 2) and two old (3 and 4) subjects (SUBJ). Total and phospho-ERK standard proteins (STD) are shown on blots. Data are means ± SEM from 12 old and 12 young subjects. *p=0.041 old skin versus young skin for phospho-ERK. (b) ERK activity was measured in soluble skin extracts, from young (mean age 23.8 y, N=15) and old (mean age 84.3 y, N=15) persons, using myelin basic protein as the substrate. Insets show representative bands of phosphorylated myelin basic protein from two young (1 and 2) and two old (3 and 4) subjects (SUBJ). Data are means ± SEM from 15 old and 15 young subjects. *p=0.05 old skin versus young skin. (c) Cyclin D2 expression was measured in soluble skin extracts from young (mean age 23.6 y, N=12) and old (mean age 83.8 y, N=11) persons by Western blot. Inserts show representative Western blots from two young (1 and 2) and two old (3 and 4) subjects (SUBJ). Data are means ± SEM from 11 old and 12 young subjects. *p=0.05 old skin versus young skin.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Stress-activated MAP kinase pathway is increased in old human skin in vivo. (a) Total JNK1 and JNK2 protein levels were determined in soluble skin extracts from young (mean age 23.6 y, N=12) and old (mean age 84.2 y, N=12) persons by Western blot. Inserts show representative Western blots from two young (1 and 2) and three old (3 and 4) subjects (SUBJ). Data are means ± SEM from 12 old and 12 young subjects. (b) Total p38 protein levels were determined in soluble skin extracts from young (mean age 23.6 y, N=12) and old (mean age 84.2 y, N=12) persons by Western blot. Insets show representative Western blots from two young (1 and 2) and two old (3 and 4) subjects (SUBJ). Data are means ± SEM from 12 old and 12 young subjects. *p=0.049 old skin versus young skin. (c) c-Jun kinase activity was determined in soluble skin extracts from young (mean age 23.8 y, N=12) and old (mean age 84.3 y, N=12) persons, by solid phase assay using GST-c-Jun as substrate. Inserts show representative phosphorylated GST-c-Jun bands from two young (1 and 2) and two old (3 and 4) subjects (SUBJ). Data are means ± SEM from 12 old and 12 young subjects. *p=0.027 old skin versus young skin.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
c-Jun, but not c-Fos, is elevated in old human skin in vivo. (a) c-Jun and c-Fos mRNA expression in skin from young (mean age 23.4 y, N=7) and old (mean age 83.3 y, N=9) persons was measured by semi-quantitative RT-PCR. Total RNA was isolated and reverse transcribed as described in Materials and Methods. The cDNA was used as a template for PCR amplification using c-Jun, c-Fos, and 36B4-specific primers. Reaction products were separated by agarose gel electrophoresis, stained with Vistra Green and quantified by fluorescence intensity. Bar heights are means ± SEM from nine old and seven young subjects. *p=0.012 old skin versus young skin for c-Jun mRNA expression. (b) Immunohistologic localization of c-Jun and c-Fos proteins in young and old skin. Photomicrographs are representative of six young (mean age 26 y) and six old (mean age 85 y) persons. Scale bar: 10 μm. Area in insets are 2-fold enlargements.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Retinol treatment stimulates ERK in old human skin in vivo. Buttock skin of old persons was treated with 1% retinol or its vehicle for 7 d. Soluble skin extracts were prepared and analyzed for ERK protein and activity. (a) Total (▪) and phosphorylated (□) ERK protein was determined in soluble extracts from vehicle-treated (V) and retinol-treated (ROL) skin of old persons (mean age 83.8 y, N=11), by Western blot. Insets show representative Western blots from two (1 and 2) subjects (SUBJ). Total and phospho-ERK standard proteins (STD) are shown on blots. Data are means ± SEM from 11 subjects. *p=0.22 retinol-treated versus vehicle-treated skin for phospho-ERK. (b) ERK activity was determined in soluble extracts from vehicle-treated (V) and retinol-treated (ROL) skin of old persons (mean age 83.1 y, N=10), using myelin basic protein as a substrate. Insets show representative bands of phosphorylated myelin basic protein from two (1 and 2) subjects (SUBJ). Data are means ± SEM from 10 subjects. *p=0.005 retinol-treated versus vehicle-treated skin.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions