Figures
- Fig. 1—
Classification of samples from groups 2 and 3, based on microRNAs differentially expressed between these groups. Dendrogram for clustering experiments, using centred correlation and average linkage. The dendrogram, based on the differentially expressed microRNAs between groups 2 (metaplasia, mild and moderate dysplasia) and 3 (severe dysplasia, carcinoma in situ (CIS) and invasive carcinoma), segregated samples of groups 2 and 3 into two groups. Only three samples of group 3 (two severe dysplasias and one CIS) were misclassified into group 2.
- Fig. 2—
Classification of samples from groups 2 and 3A (A), based on microRNAs differentially expressed between these groups. Dendrogram for clustering experiments, using centred correlation and average linkage. The dendrogram, based on the differentially expressed microRNAs between groups 2 (metaplasia, mild and moderate dysplasia) and 3A (severe dysplasia and carcinoma in situ (CIS)), segregated samples of groups 2 and 3A into two groups. Only three samples of group 3A (two severe dysplasias and one CIS) were misclassified into group 2.
- Fig. 3—
Classification of samples between carcinoma in situ (CIS; stage 8) and invasive squamous cell carcinoma (SQCC; stage 9), based on microRNAs differentially expressed between these groups. a) Dendrogram for clustering experiments, using centred correlation and average linkage. MicroRNAs differentially expressed between CIS and SQCC segregated the samples of these stages into two groups according to their histological classification: the six samples of stage 8 and of stage 9 were well separated into two different branches. b) The log intensities are directly proportional to the cycle threshold (CT), and the green (smaller CT) and the red (higher CT) represent up- and downregulated microRNAs, respectively. The first five microRNAs were upregulated in SQCC (the first six samples) compared with CIS (the last six samples). The last five microRNAs were downregulated from CIS to SQCC.
Tables
- Table 1—
Composition of groups
Nonsmokers Smokers Nl ns Nl Nfl Nl Hf H M MiD MoD SD CIS SQCC Stage 0 1 2 3 4 5 6 7 8 9 Group 0 1 1 1 2 2 2 3A# 3A# 3B# Nl ns: normal of nonsmokers; Nl Nfl: normal in normofluorescent area; Nl Hf: normal in hypofluorescent area; H: hyperplasia; M: metaplasia; MiD: mild dysplasia; MoD: moderate dysplasia; SD: severe dysplasia; CIS: carcinoma in situ; SQCC: squamous cell carcinoma. Stages 0 to 9 were assembled in larger groups of samples according to transcriptomic analyses performed on the same samples, thereby increasing the power of statistical tests. #: subdivision of group 3.
- Table 2—
Statistically significantly evolving microRNAs during lung carcinogenesis
MicroRNA name Groups compared 0 and 1 0 and 2 0 and 3 1 and 2 2 and 3 2 and 3A 3A and 3B CIS and 3B hsa-miR-1 Down 5.88 Down 10.00 hsa-miR-7 Up 4.11 Up 3.96 hsa-miR-9 Down 11.11 Up 17.89# Up 15.97# Up 3.05 hsa-miR-9* Up 2.49 Up 5.66 hsa-miR-10b Down 3.03 Up 2.95 hsa-miR-15a Down 5.00 Down 6.67 Down 3.45 hsa-miR-17-5p Up 2.15 hsa-miR-32 Down 6.67 Down 5.88 Down 4.17 hsa-miR-34a Down 5.00 hsa-miR-34c Down 6.67 Down 14.29# Down 25.00# Down 2.50 hsa-miR-99a Down 1.96 Down 1.89 hsa-miR-101 Down 5.56 Down 3.57# hsa-miR-106b Up 2.06 Up 2.36 hsa-miR-132 Up 2.18 hsa-miR-133a Down 5.88 hsa-miR-134 Down 2.70 hsa-miR-135a Down 4.17 Down 4.00# Down 4.00 hsa-miR-139 Down 2.27 Down 5.26 Down 5.88# hsa-miR-142-3p Down 2.94 Down 5.56 Up 2.60 Up 2.50 Up 2.51 Up 2.24 hsa-miR-142-5p Down 2.38 Up 2.86 Up 3.36 hsa-miR-143 Down 3.85 hsa-miR-145 Down 2.70 has-miR-183 Up 2.66 hsa-miR-190 Down 4.76 hsa-miR-193a Down 8.33# hsa-miR-193b Down 4.17 Up 5.66 hsa-miR-196a Up 16.95# Up 25.69# Up 28.29# hsa-miR-196b Up 10.48 Up 10.80# Up 10.78# hsa-miR-199a Up 8.77# Up 20.87 hsa-miR-203 Up 5.26# Up 4.89 Up 5.79# hsa-miR-204 Down 11.11 Down 16.67 hsa-miR-206 Up 3.86 Down 2.33 hsa-miR-213 Down 3.57 hsa-miR-214 Down 2.00# Up 2.35 Up 2.67 hsa-miR-216 Down 3.33 Down 2.63 Down 2.44 hsa-miR-218 Down 5.56 Down 3.57 hsa-miR-224 Up 2.58 Up 1.63# Up 3.79# Up 1.63 hsa-miR-301 Down 3.57 Up 2.47 hsa-miR-324-5p Down 10.00 hsa-miR-338 Up 5.07 hsa-miR-361 Down 6.25 Down 4.76 hsa-miR-362 Up 2.75 Up 3.36 hsa-miR-374 Down 1.49 hsa-miR-375 Down 6.25 Down 7.14# Down 11.11 hsa-miR-382 Down 2.44 hsa-miR-410 Down 2.63 hsa-miR-411 Down 4.76 hsa-miR-422b Down 4.76 Down 4.76 Down 4.17 hsa-miR-432 Up 3.46 hsa-miR-449 Down 3.33 Down 5.56 Down 5.56 hsa-miR-449b Down 11.11 hsa-miR-451 Down 8.33 hsa-miR-452 Up 14.81# Up 6.47# Up 6.10# hsa-miR-486 Down 10.00 hsa-miR-487b Up 8.19 Up 19.03# hsa-miR-492 Down 2.5 hsa-miR-518d Down 2.63 hsa-miR-521 Down 1.56 hsa-miR-545 Down 2.78 hsa-miR-565 Down 1.67 Down 2.08 hsa-miR-572 Down 3.45 Down 3.85 hsa-miR-597 Up 5.43 hsa-miR-615 Up 9.74 hsa-miR-622 Down 6.25 hsa-miR-639 Down 2.86 has-miR-642 Down 5.00# Down 12.50 Down 2.17 Down 3.33 Down 6.67 hsa-miR-646 Down 5.00 hsa-miR-650 Up 4.97 hsa-miR-659 Up 4.83 Down 3.70 Up 6.81 Data are presented as fold changes. CIS: carcinoma in situ; down: downregulated; up: upregulated. All the microRNAs found differentially expressed in at least one of the eight comparisons are shown (t-test p<0.05). #: microRNAs remaining statistically differentially expressed in a specific comparison after correction for multiple testing (CMT) at Bonferroni corrected p-value <0.05. In the first comparison (group 0 versus group 1), no microRNA expression difference remained statistically significant after CMT, whereas, in the seven other comparisons, the numbers of microRNAs with differential expression remaining statistically significant after CMT were three, six, four, five, four, one and two (from left to right).
- Table 3—
Two-step evolution of microRNAs in lung squamous carcinogenesis
Differentially expressed microRNAs Groups compared 0 and 1 0 and 2 0 and 3 1 and 2 2 and 3 2 and 3A 3A and 3B CIS and 3B Total number 6 19 19 16 30 15 20 10 Downregulated 5 (83) 16 (84) 14 (74) 14 (87.5) 17 (57) 3 (20) 8 (40) 5 (50) Upregulated 1 (17) 3 (16) 5 (26) 4 (12.5) 13 (43) 12 (80) 12 (60) 5 (50) Data are presented as n or n (%). CIS: carcinoma in situ. Two successive waves were observed in microRNA expression profiles during lung squamous carcinogenesis: first, a downregulation and, secondly, an upregulation of microRNAs. In the first four comparisons of microRNA profiles, between groups 0 and 1, 0 and 2, 0 and 3, and 1 and 2, the large majority of microRNAs were downregulated. In the later four comparisons, the proportion of downregulated microRNAs decreased and that of upregulated microRNAs increased, with ≥50% being upregulated for comparisons between groups 2 and 3A, 3A and 3B, and CIS and 3B.
