Table 3

Interindividual differences in CYP1B1 and GSTP1 expression in buccal cells from 19 active smokers

Subject IDPlasma nicotine (ng/ml)Plasma cotinine (ng/ml)CYP1B1 qualitative expression (+ or −)Quantitative Expression: CYP1B1 CRO scaled to ref. transcript CRO, same RNA/cDNA samplePt:CYP1B1 scaled to mcf7:CYP1B1GSTP1 qualitative expression (+ or−)Quantitative expression: GSTP1 CRO scaled to ref. transcript CRO, same RNA/cDNA samplePt:GSTP1 scaled to Cl-GSTP1 (PtGSTP1)− (Cl-GSTP1)
(PtCYP1B1)GAPDH)Pt(CYP1B1BAUP2(PtCYP1B1)(ClCYP1B1)Pt(GSTP1GAPDH)Pt(GSTP1BAUP2)
15315.00199+2.907.7813.61nananana
16218.30202nana12.45+nana14.3
1680019.7724.9627.50+10.5813.789.79
17116.5206+1.466.8514.48+nanana
17217.1217nanana+nanana
181069.8+nana10.09+2.357.8611.6
18811.2276+1.02na11.2614.9515.1821.54
198nana+2.867.4211.46+na7.178.1
20517.3256+2.104.1013.00+12.211.4118.36
20616.4228+6.958.1914.63+10.199.612.24
20919.2134+9.4111.5512.28+3.37na5.75
2105.4119+-1.483.5011.28+7.1812.1610.99
21317.7276+0.86na7.93+9.8110.337.11
21615.9186+nana13.44+nana7.32
2185.7410915.5217.4328.75+1213.8512.97
22728.8210+3.808.4217.19+nanana
2321.58113+7.9013.2914.94+10.9816.218.53
50440+na0.6612.3218.220.425.96
50491.77110+nananananana
  • NOTE: Example of a data set for quantitation of CYP1B1 and GSTP1 transcript levels in brushed exfoliated buccal cells from the 19 current cigarette smokers. The method employed RNA-specific (18) real-time quantitative RT-PCR. Values represent LightCycler (Roche) thermocycler-generated CRO differences between the target gene transcript and the reference target transcript. For aid with interpretation, 1/[2 (CRO:T) − (CRO:Ref)] represents the ratio of target (T) to reference transcript (R). Therefore, the smaller the CRO difference, the higher the amount of target transcript is present relative to the reference transcript. For example, in column five, for CYP1B1 levels normalized to GAPDH in two subjects (205 and 206) with virtually identical tobacco exposure, as measured by plasma nicotine and cotinine, subject 206 expresses 1/2[6.95−2.10] = 1/2 [4.85], or only 3.5% of the amount of CYP1B1 expressed by subject 205; similar results were found when β-actin was used as the referent transcript. “Cl” and “MCF-7”denote reference pool of total human RNA either extracted from commercially obtained lung tissue RNA (Clontech) or from MCF-7 breast cancer cells exposed to dioxin, respectively; these served as external reference control samples. Absent transcript signal after the 50 PCR cycle protocol was assigned an arbitrary CRO value of 46, the limit of detection of the Roche LightCycler system, as stated in Materials and Methods.

  • Abbreviations: CRO, crossover threshold; na, samples unavailable for quantitation.