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Validation of a Novel Fgf10 ^Cre-ERT2 Knock-in Mouse Line Targeting FGF10^Pos Cells Postnatally.

ABSTRACT: Fgf10 is a key gene during development, homeostasis and repair after injury. We previously reported a knock-in Fgf10 ^Cre-ERT2 line (with the Cre-ERT2 cassette inserted in frame with the start codon of exon 1), called thereafter Fgf10 ^Ki-v1, to target FGF10^Pos cells. While this line allowed fairly efficient and specific labeling of FGF10^Pos cells during the embryonic stage, it failed to target these cells after birth, particularly in the postnatal lung, which has been the focus of our research. We report here the generation and validation of a new knock-in Fgf10 ^Cre-ERT2 line (called thereafter Fgf10 ^Ki-v2) with the insertion of the expression cassette in frame with the stop codon of exon 3. Fgf10 ^Ki-v2/+ heterozygous mice exhibited comparable Fgf10 expression levels to wild type animals. However, a mismatch between Fgf10 and Cre expression levels was observed in Fgf10 ^Ki-v2/+ lungs. In addition, lung and limb agenesis were observed in homozygous embryos suggesting a loss of Fgf10 functional allele in Fgf10 ^Ki-v2 mice. Bioinformatic analysis shows that the 3'UTR, where the Cre-ERT2 cassette is inserted, contains numerous putative transcription factor binding sites. By crossing this line with tdTomato reporter line, we demonstrated that tdTomato expression faithfully recapitulated Fgf10 expression during development. Importantly, Fgf10 ^Ki-v2 mouse is capable of significantly targeting FGF10^Pos cells in the adult lung. Therefore, despite the aforementioned limitations, this new Fgf10 ^Ki-v2 line opens the way for future mechanistic experiments involving the postnatal lung.

SUBMITTER: Chu X

PROVIDER: S-EPMC8155496 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundPrognostic biomarkers of treatment response to distinct biologic disease-modifying anti-rheumatic drugs (b-DMARDs) are still lacking within the management of rheumatoid arthritis (RA).MethodsThirty-four b-DMARDs naive RA patients, divided by disease duration into early (cohort 1) and long standing (cohort 2), received CTLA4-Ig. At study entry, and every 3 months for 1 year, each patient underwent peripheral blood (PB)-derived CD4pos cell subpopulation assessment by flow cytometry, STAT3 and STAT5 expression by RT-PCR and IL-6, IL-12p70, TGFβ, and IL-10 serum levels by ELISA. The DAS and CDAI remission was assessed at 6 and 12 months.ResultsDAS- and CDAI-defined remission within 12 months was achieved by 16 (47.1%) and 8 (23.5%) RA patients, respectively. Considering the whole RA cohort, CTLA4-Ig induced a significant decrease of IL-6 serum levels from baseline to 6 and 12 months, as well as of PB CD4posCD25posFoxP3pos cells at 6 and 12 months, and of CD4posIL17pos cells after 12 months. PB CD4pos cells of RA patients showed higher STAT3 and STAT5 expression than healthy controls, which remained unchanged within 12 months of treatment. At study entry, RA patients achieving DAS remission had significantly lower IL-6 serum levels than RA patients not achieving this outcome. In particular, having baseline IL-6 serum levels ≤ 8.4 pg/ml, significantly identified naïve to b-DMARDs RA patients more likely to achieve DAS-remission under CTLA4-Ig at 6 months (66.7%) compared to RA patients with baseline IL-6 serum levels > 8.4 pg/ml [15.4%, OR (95%Cis) 11.00 (1.75-55.82)]. Moreover, having CD4posCD25posFoxP3pos cells rate ≥ 6.0% significantly identifies naïve to b-DMARDs early RA patients more likely to achieve DAS remission at 6 months (83.3%) compared to RA patients with baseline CD4posCD25posFoxP3pos cells < 6.0% [16.7%, OR (95% Cis) 25.00 (1.00-336.81)].ConclusionsBaseline IL-6 serum levels and peripheral blood-derived CD4pos subpopulations are putative novel prognostic biomarkers of CTLA4-Ig response in RA patients.

Project description:BackgroundEvidence on the changes in the absolute counts of monocyte subpopulations in sepsis is missing.MethodsFirstly, absolute counts of circulating CD14pos/HLA-DRpos/CD45pos monocytes were measured by flow cytometry in 70 patients with Gram-negative sepsis and in 10 healthy volunteers. In the second phase, immunophenotyping was performed and the absolute count of circulating inflammatory monocytes and of circulating CD14dim/CD16pos/CD45pos patrolling monocytes were measured in another 55 patients and 10 healthy volunteers. Measurements were repeated on days 3, 7, and 10. Results were correlated with survival after 28 days.ResultsGreater numbers of CD14pos/HLA-DRpos/CD45pos monocytes were found on day 1 in survivors compared to nonsurvivors (p = 0.030). Receiver operating characteristic (ROC) analysis showed that a cutoff higher than 337 cells/mm3 on day 1 could discriminate between survivors and nonsurvivors with a positive predictive value (PPV) of 91.1%. Logistic regression including Sequential Organ Failure Assessment (SOFA) score and Acute Physiology and Chronic Health Evaluation (APACHE) II score showed that an absolute count greater than 337 cells/mm3 was independently associated with unfavorable outcome (odds ratio (OR) 0.19, p = 0.050). The absolute counts of inflammatory and of CD14dim/CD16pos/CD45pos monocytes were greater in patients than healthy controls during the entire 10 days of follow-up. The absolute counts on day 3 of CD14dim/CD16pos/CD45pos monocytes were greater in survivors than nonsurvivors (p = 0.027). ROC analysis revealed that the cutoff at 27 cells/mm3 could discriminate between survivors and nonsurvivors with PPV of 94.1%. Logistic regression including age, SOFA score, and APACHE II score showed that an absolute count greater than 27 cells/mm3 was independently associated with unfavorable outcome (OR 0.06, p = 0.033). Logistic regression analysis showed that intra-abdominal infection on day 1 was predictive of low CD14dim/ CD16pos/CD45pos count on day 3.ConclusionCirculating counts of inflammatory and patrolling monocytes are greatly increased in Gram-negative sepsis. Absolute counts of CD14pos/HLA-DRpos/CD45pos monocytes on day 1 and CD14dim/CD16pos/CD45pos monocytes on day 3 are independently associated with final outcome.Trial registrationClinicalTrials.gov, NCT01223690 . Registered retrospectively on 18 October 2010.

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Validation of a Novel Fgf10 ^Cre-ERT2 Knock-in Mouse Line Targeting FGF10^Pos Cells Postnatally.

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Dataset Information

Validation of a Novel Fgf10 Cre-ERT2 Knock-in Mouse Line Targeting FGF10Pos Cells Postnatally.

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OmicsDI is part of the ELIXIR infrastructure

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Validation of a Novel Fgf10 ^Cre-ERT2 Knock-in Mouse Line Targeting FGF10^Pos Cells Postnatally.