Induction of Collagenase mRNA Expression in Dermal Fibroblasts by IFN- <i>α</i> 2b and Determination of the IFN- <i>α</i> 2b Responsive Element on 5′-Flanking Regions of Collagenase Promoter
Bibliographic record
Abstract
We have previously demonstrated that interferon-alpha2b (IFN-alpha2b) markedly depresses the expression of mRNA for type I procollagen in dermal fibroblasts. In the present study, the effect of various concentrations of IFN-alpha2b on the expression of collagenase mRNA and activity of 5'-flanking regions of collagenase promoter in dermal fibroblasts are presented. The results showed at least a 2-fold increase in the expression of collagenase mRNA in fibroblasts grown at either 70% confluency (40.9 +/- 4.6 vs. 18.5 +/- 1.6, n=4, p<0.05) or 95% confluency (24.7 +/- 6.7 vs. 4.5 +/- 1.6, n=4, p<0.05). The effects of IFN-alpha2b on collagenase mRNA stability and promoter activity were evaluated to determine the mechanism by which IFN-alpha2b increases the expression of collagenase mRNA. IFN-alpha2b-treated and untreated fibroblasts were treated with alpha-amanitin to arrest collagenase mRNA transcription, and total RNA was then harvested at 0, 3, 6, 12, and 24 h. The decay curves of collagenase mRNA as a function of time showed a greater rate of degradation for collagenase mRNA in IFN-alpha2b-treated cells relative to untreated control cells. This difference was more pronounced in cells treated with alpha-amanitin at either 12 or 24 h. To determine the regions of the collagenase promoter that might function as IFN-alpha2b responsive elements, eight different fragments of the collagenase promoter, -518, -300, -171, -161, -127, -91, -74, and -66 to +63 nucleotide (nt), were constructed in a chloramphenicol acetyltransferase (CAT) expression vector. The results of CAT activity of cells transfected with these construct identified three constructs, 171/+63, -161/+63, and -127/+63, as being responsive to IFN-alpha2b treatment in dermal fibroblasts. The CAT activity was increased 279%, 163%, and 261% in -171/+63, -161/+63, and -127/+63-transfected fibroblasts, respectively, in response to IFN-alpha2b treatment relative to untreated control. No significant increase in CAT activity was found in cells transfected with the other constructs of the collagenase promoter. A time response experiment showed a marked increase in CAT activity of cells transfected with either 127/+63 or -171/63 constructs within 6-12 hr of IFN-alpha2b treatment. In conclusion, IFN-alpha2b significantly increases the expression of collagenase mRNA in dermal fibroblasts probably through stimulation of the -127/-91 region of the collagenase promoter. Thus, this region may function as an IFN-alpha2b responsive element on collagenase promoter.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.002 | 0.001 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".