Cold-Induced Lipoprotein Clearance in Cyp7b1-Deficient Mice
The main results reproduced: recomputed values matched the published ones within tolerance.
Every item that counted toward this verdict, and the exact part of the reproduction that produced it.
- ✓Same input data as the authors
- ✓Reported values were directly comparable
- ✓No relevant deviation in data/preprocessing
- ✓No authors-side cause for any deviation
- ✓Reported values are derivable from the shared data
- ✓Any deviation was negligible
- ✓The central claim held under reproduction
- ✓Overall, the reproduction was clean
- Every checked point held up.
A 0–100 reproducibility-quality score from the per-question grades, shown as a z-score: standard deviations above (+) or below (−) the mean of comparable assessments.
▸Reproduction agent’s raw note
Wet-lab metabolic-physiology paper; NO bioinformatic pipeline, no GEO/SRA, no code. Only deposited computational data = a small custom TaqMan qPCR panel on Mendeley (doi:10.17632/ywcpv6wwzr.1, 18 genes x 28 samples). In scope = the qPCR-derived gene-expression figures (1B liver, 5A BAT); reproduced by re-deriving fold-change and group statistics. RESULT = partial-but-strong: Fig 5A (BAT, WT-cold n=9 vs KO-cold n=8) reproduces cleanly - all 14 genes match the reported direction, with Lpl-down, Lipg-down and Angptl4-up all significant (p<0.05) exactly as reported, 'unchanged' genes ns, and all six 'trend-up' lipogenesis genes up. Fig 1B headline Cyp7b1 result reproduces exactly from the normalized matrix (cold induction in WT 2.82x p=0.008; knockout-low 0.20-0.51x p=0.008). Computed locally (no heavy compute needed; 58 KB of tables). Ran no «our HPC» job. NOT attempted (out of scope): all radiotracer turnover, plasma/insulin/bile-acid assays, western blots, ELISA, HOMA-IR. KEY FLAGS: (1) the deposited LIVER 'Fold Change' worksheet is internally inconsistent - not derivable from the normalized matrix and shows KO Cyp7b1 ~1.8x (high), contradicting the knockout; the normalized matrix is correct and was used instead (flag for human review). (2) Housekeeping Tbp Ct not deposited -> Ct->normalized step uncheckable. (3) 2 of 20 panel genes (Slc2a1/Slc2a4) not deposited.
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Assessment versions
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v1 current initial assessment Score 83assessed: 2026-06-18 ⛓ 05ac4a26dfcd
✎ I am an author of this paper
Updated or fixed a deposit, or is there an erratum? Ask us to re-run the metrics. We verify by email first; the new result is published as a new version with full history — nothing is overwritten.
Provenance — full disclosure
When this reproduction was carried out, which methodology version was used, and by whom — so the record can be audited and checked independently.
- Reproduced
- 2026-06-18
- Rubric version
- v1.0
- Assessed by
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🤖 AI curator · v1.0 · run #1 2026-06-18no human curator yet
- Last updated
- 2026-08-05
Provisional, curator- or AI-assessed, and independently checkable. A reproduction outcome states what one attempt could reproduce — not a judgement of the authors.
Deep full-text extraction
Model: sonnetThe study tests whether loss of Cyp7b1 (and the resulting reduction in cold-induced bile acid synthesis) impairs BAT-mediated systemic postprandial lipoprotein metabolism when brown adipose tissue is activated by cold exposure.
- ★ Cyp7b1 deficiency blunts cold-induced hepatic Cyp7b1 upregulation and plasma bile acid synthesis finding
- ★ Cyp7b1 deficiency compromises BAT thermogenic activation (reduced pPKA substrates, T3, and OXPHOS complex I/II) in response to cold finding
- ★ Cyp7b1-/- mice show diminished cold-induced BAT uptake of triolein-derived fatty acids (3H) and lipoprotein-core cholesterol remnants (14C) after an oral fat tolerance test finding
- ★ Impaired BAT lipid uptake in Cyp7b1-/- mice is linked to reduced BAT lipoprotein lipase (LPL) levels and compromised organ activity, possibly related to impaired insulin signaling mechanism
- Bile acids activate BAT thermogenesis via the TGR5/PKA pathway, increasing Dio2 expression mechanism
- ★ Cold housing increases plasma bile acid levels across all BA species in WT mice, an effect reversed in Cyp7b1-/- mice finding
- Cold exposure lowers plasma triglycerides and total cholesterol irrespective of genotype finding
| Assay | System | Perturbation | Readout | Platform |
|---|---|---|---|---|
| qRT-PCR gene expression | liver, WT and Cyp7b1-/- mice | Cyp7b1 KO + cold (6°C) vs thermoneutral (30°C) housing | mRNA levels of BA synthesis genes (Cyp7b1, Cyp27a1, Cyp7a1, Cyp8b1) | ABI 7900HT / QuantStudio 5 with TaqMan primers |
| Western blot | liver, WT and Cyp7b1-/- mice | Cyp7b1 KO + cold vs thermoneutral | CYP7B1 protein levels | Amersham Imager 600 / ECL |
| Bile acid quantification (HPLC-ESI-MS/MS, MRM) | plasma, WT and Cyp7b1-/- mice | Cyp7b1 KO + cold vs thermoneutral | circulating bile acid species levels | Shimadzu Nexera X2 HPLC + SCIEX QTrap 5500 QqQ |
| Western blot | interscapular BAT (iBAT), WT and Cyp7b1-/- mice | Cyp7b1 KO + cold housing | OXPHOS complexes I-V and phosphorylated PKA substrates | Amersham Imager 600 / ECL |
| T3 ELISA | BAT tissue extract, WT and Cyp7b1-/- mice | Cyp7b1 KO + cold housing | triiodothyronine (T3) levels | CALBIOTECH T3043T-100 kit |
| Oral fat tolerance test with 3H-triolein/14C-cholesterol radiotracers | whole animal (plasma, iBAT, ingWAT, gonWAT, heart, liver, spleen, kidney, muscle), WT and Cyp7b1-/- mice | Cyp7b1 KO + cold vs thermoneutral, oral corn oil gavage | blood and organ radioactivity (fatty acid and cholesterol uptake) | Perkin Elmer Tricarb Scintillation Counter |
| Plasma triglyceride assay | plasma, WT and Cyp7b1-/- mice | cold vs thermoneutral, postprandial (OFTT) | plasma TG concentration over 120 min | Roche colorimetric kit, 96-well format |
| Plasma insulin ELISA | plasma, WT and Cyp7b1-/- mice | cold vs thermoneutral | insulin levels | Chrystal Chem #90060 ELISA kit |
- ▲ Hepatic Cyp7b1 expression and protein strongly upregulated by cold in WT but near-absent in Cyp7b1-/- mice at both temperatures
- ▲ Plasma BA levels increased with cold in WT mice, blunted in Cyp7b1-/- mice
- – iBAT uptake of 3H-triolein-derived fatty acids increased with cold in WT, significantly diminished (but not abolished) in Cyp7b1-/- mice
- ▼ iBAT uptake of 14C-cholesterol increased with cold in both genotypes but reduced in cold-housed Cyp7b1-/- vs WT
- ▼ Plasma TG rose only moderately in cold-housed mice vs doubling in warm-housed mice, independent of genotype; TG lower after 120 min in cold groups
- ▼ OXPHOS complex I and II protein abundance significantly lower in cold-housed Cyp7b1-/- BAT; complexes III-V unaffected
- ▼ pPKA substrate levels (mainly ~40 kDa proteins) reduced in BAT of cold-housed Cyp7b1-/- mice; CREB phosphorylation reduced by trend
- ▼ T3 levels lower in BAT of cold-housed Cyp7b1-/- mice
- count n = 3-4/group (hepatic gene expression and plasma BA levels, Figure 1B/E)
- count n = 9 (WT), n = 8 (Cyp7b1-/-) (Western blot OXPHOS/pPKA quantification, cold-exposed mice, Figure 1F/G)
- count n = 5/group (oral fat tolerance test radiotracer study, Figures 2 and 3)
- pvalue p < 0.05 (significance threshold for ANOVA/t-test group comparisons (ABC letter system))
- other 3H-triolein 185 kBq/mouse, 14C-cholesterol 37 kBq/mouse (radiotracer dosing in oral fat tolerance test)
- other Triton WR-1339, 5 g/kg body weight (chylomicron production assay tail-vein injection dose)
Statistical methods review
Model: sonnetA neutral, descriptive read of the statistical approach — what was done, and (for shared learning, not as criticism) what could also have been done.
This study used a 2×2 factorial design (genotype: Cyp7b1−/− vs wild-type littermates × housing temperature: 30°C vs 6°C) in male mice to investigate cold-induced lipoprotein clearance. Group comparisons employed Student's t-test and one- or two-way ANOVA with Tukey post-hoc correction, with significance set at p<0.05 indicated via an ABC letter system. Results are consistently presented as mean ± SEM, with GraphPad Prism 7.0 used for all statistical calculations.
| Test | Applied to | n | Assumptions |
|---|---|---|---|
| Two-way ANOVA with Tukey post-hoc correction | Figure 1G: OXPHOS complex and pPKA substrate protein levels in cold-exposed WT vs Cyp7b1−/− mice; explicitly stated in figure legend; also applied to other genotype × temperature comparisons per Methods | WT n=9, Cyp7b1−/− n=8 (cold-exposed groups, Figure 1G); n=3–4/group (Figure 1E plasma BA) | not stated |
| One-way ANOVA with Tukey post-hoc correction | Unspecified comparisons across figures (stated in Methods as used alongside two-way ANOVA and t-test) | — | not stated |
| Student's t-test (direction not specified) | Unspecified pairwise comparisons across figures (stated in Methods) | — | not stated |
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Dispersion is reported as SEM throughout, with group sizes of n=3–9↳ Could also: Standard deviation (SD) or 95% confidence intervals could also be used to express variability — SD directly describes within-group biological spread regardless of n; 95% CIs additionally convey estimation precision; both are widely recommended over SEM for small-sample animal studies because SEM shrinks with n and can give the impression of tighter data than SD would show
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Student's t-test was used alongside ANOVA for group comparisons, though the allocation of specific contrasts to each test is not described↳ Could also: All pairwise and factorial contrasts could be unified within the ANOVA framework using Tukey or Dunnett post-hoc tests — A single ANOVA-based framework consistently controls the family-wise error rate across all comparisons and avoids ambiguity about which contrasts were tested with which method
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Parametric tests (t-test, ANOVA) were applied with group sizes of n=3–5 in several experiments↳ Could also: Non-parametric alternatives such as Mann-Whitney U (pairwise) or Kruskal-Wallis with Dunn post-hoc (multi-group) could also be applied — With very small n, distributional assumptions underlying parametric tests are difficult to verify empirically; non-parametric tests make no normality assumption and are a common alternative in small-sample preclinical studies
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Significance is reported via binary p<0.05 letter notation without exact p-values↳ Could also: Exact p-values and effect size measures (e.g., Cohen's d, partial η²) could also be reported — Exact p-values convey graduated strength of evidence beyond a threshold, and effect sizes allow assessment of biological magnitude independently of sample size; both are increasingly required by reporting guidelines (e.g., Nature journals, APA) and facilitate meta-analysis
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The oral fat tolerance test includes serial blood draws at 0, 60, and 120 min, analyzed with ANOVA across groups↳ Could also: A repeated-measures ANOVA or linear mixed-effects model with time as a within-subject factor could also be used for the time-course radiotracer data — Because repeated measurements from the same animal are correlated, a model that explicitly accounts for within-subject structure uses the data more efficiently and correctly partitions variance between time, genotype, temperature, and their interactions
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Radiotracer uptake values (organ-level counts) were analyzed with ANOVA on the original scale↳ Could also: Log-transformation of count-based radioactivity data prior to ANOVA could also be applied — Radioactivity count data often exhibit right-skewed distributions and variance that scales with the mean; log-transformation can stabilize variance and better satisfy ANOVA homoscedasticity and normality assumptions, particularly when group means differ substantially
What was reproduced
The exact results taken into scope, with each reported value next to the value our attempt produced.
Scope — PMID 35478959
Title: Cold-Induced Lipoprotein Clearance in Cyp7b1-Deficient Mice Journal: Front Cell Dev Biol 2022 · DOI 10.3389/fcell.2022.836741 · PMC9038073 Authors' data: Mendeley Data doi:10.17632/ywcpv6wwzr.1 ("Gene Expression Liver/BAT Cyp7b1ko warm cold") No GEO / no SRA / no code repository (verified via elink pubmed→gds/sra: none).
This is a wet-lab metabolic-physiology study. There is no bioinformatic pipeline (no sequencing, no omics). The only deposited "raw data" is a small custom TaqMan qPCR gene-expression panel (4 xlsx tables on Mendeley).
IN SCOPE (pipeline/computation-derived, reproducible from deposited data)
The qPCR-derived gene-expression panels and their downstream computation (housekeeping normalization → fold-change → group statistics):
- Figure 1B — hepatic bile-acid-synthesis genes: Cyp27a1, Cyp7a1, Cyp7b1, Cyp8b1 (liver, 4 groups: WT/KO × warm/cold; n=2–3/group).
- Figure 5A — BAT lipoprotein-processing / lipogenesis genes: Lpl, Lipg, Gpihbp1, Angptl4, Pnpla2, Lipe, Cd36, Slc27a1/4/5, Chrebp-beta, Fasn, Acaca, Scd1 (BAT, WT cold n=9 vs KO cold n=8).
Pipeline = qPCR 2^-ΔΔCt / Tbp-normalized copy numbers → fold change vs control group → two-way ANOVA (liver) / group comparison (BAT). GraphPad Prism 7.0.
OUT OF SCOPE (wet-lab / manual / external — not attempted)
- Metabolic turnover studies with ³H-triolein/¹⁴C-cholesterol radiotracers, scintillation counting (Figs 2, 3, 4) — wet-lab.
- Plasma TG / cholesterol / insulin kits, bile-acid LC-MS/MS (Fig 1E, Suppl S1), T3 ELISA (Suppl S2B) — wet-lab assays.
- Western blots + densitometry (CYP7B1, OXPHOS, pPKA, LPL, GPIHBP1, FASN, pAKT; Figs 1C/D/F/G, 5B/C/E) — wet-lab.
- HOMA-IR (Fig 5F) — derived from wet-lab plasma values not deposited.
Data limitation
The housekeeping (Tbp) Ct values are not deposited, so the non-normalized→ normalized step cannot be recomputed from scratch. The fold-change step IS checkable from the deposited normalized matrix (see AUDIT.md).
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Reproduction footprint
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