Biology subjects

Skubic, C.

Publications and source records attributed to Skubic, C..

4 recordsLinked to original sources

bioRxiv2025

Stable reference genes for 24-hour circadian profiling of core clock genes in the blood of obstructive sleep apnea patients

Obstructive sleep apnea (OSA), one of the most common breathing disorders during sleep, is associated with circadian rhythm disruption, yet circadian genes expressed in blood remain largely unexplored as biomarkers. Reliable circadian gene expression analysis requires stable reference genes for qRT-PCR normalization, but no clear guidance exists for blood-based circadian studies in OSA. We evaluated the expression of 11 candidate reference genes (GAPDH, ACTB, RPL13A, PPIB, TBP, HPRT1, PPIA, SDHA, TUBB2A, UBC) in whole blood collected every 6 hours over 24 hours from 40 participants who underwent overnight respiratory polygraphy. Gene stability was assessed with RefFinder and EndoGeneAnalyzer across OSA severity levels, intra-individual variability, and time-point variability. ACTB and RPL13A consistently emerged as the most stable reference genes under all conditions. Their robustness was confirmed by personalized cosinor analysis of core clock genes (BMAL1, PER2, CRY1), showing that reliable normalization enables detection of circadian oscillations in clinical samples. While population-level analysis revealed no significant rhythmicity, individual profiles showed oscillations in 25 participants, independent of OSA severity. These findings identify ACTB and RPL13A as strong candidates for circadian transcriptomic studies in blood and provide a methodological foundation for biomarker discovery in sleep medicine, molecular medicine, and translational chronobiology.

molecular biology↗

bioRxiv2025

Hepatic management of toxic sterols after acute deletion of Cyp51 from cholesterol synthesis

Lanosterol 14-demethylase (CYP51) is an enzyme involved in cholesterol synthesis, crucial for the normal liver function. Diminished activity of CYP51 leads to metabolism associated liver disease, ending in hepatocellular carcinoma. It is still not clear which processes are most affected in the hepatocytes and how they communicate with other cells towards the progressive liver pathology. Herein we describe a new inducible, liver-specific Cyp51 knockout mouse model (iLKO), developed to study how acute disruption of cholesterol synthesis is managed in the adult liver. Doxycycline inducible deletion avoided developmental confounders of albumin-Cre models and enabled isolation of viable primary hepatocytes. iLKO hepatocytes and liver tissue showed CYP51 depletion with accumulation of lanosterol and 24,25-dihydrolanosterol, while hepatic cholesterol levels remained largely unchanged, indicating compensatory uptake and/or pathway rerouting. Histological examination and transmission electron microscopy (TEM) revealed hepatomegaly with mild portal inflammation and ductular reaction but no overt fibrosis at the studied time points. iLKO hepatocytes displayed increased nuclear lipid droplets (LD) that might be involved in adaptation to endoplasmic reticulum (ER) stress. Additionally, we discovered crystal-like inclusions, especially in Kupffer cells. MALDI-TOF MSI could not resolve their composition, but their occurrence alongside sterol overload implicates non-cholesterol sterol crystallization as a potential trigger of inflammation. In summary, iLKO is a suitable model for dissection of sterol toxicity, clearly separated from developmental effects of Cyp51 depletion. Future examinations could reveal how toxic sterol intermediates are buffered in adult liver and how they might be connected to inflammation driven pathologies, relevant also to human health.

biochemistry↗

bioRxiv2023

Sterols from cholesterol synthesis control distinct gene regulatory pathways

Sterol intermediates of cholesterol synthesis are largely dedicated to cholesterol. Here we assess how they influence downstream gene regulatory pathways by developing knockouts (KOs) for consecutive enzymes of cholesterol synthesis in human hepatoma HepG2 cells. The KO of CYP51, DHCR24, and SC5D led to the build-up of specific sterols. The shared differentially expressed genes accounted for only 9% with regards to steroid metabolism and proliferation control, with majority of pathways changed in just one KO. The CYP51 KO cells with highly elevated 24,25-dihydrolanosterol exhibited a significant increase in G2+M phase along with enhanced cancer and cell cycle pathways, likely driven by elevated LEF1 through modulation of WNT/NFKB signalling. In contrast, SC5D and DHCR24 KO cells with elevated lathosterol or desmosterol, slowed cell proliferation and promoted apoptosis with downregulated E2F, mitosis, cell cycle transition, and enriched HNF1A tumor suppressor. These findings demonstrate that sterols from cholesterol synthesis control distinct gene regulatory pathways, while only early sterols can promote cell proliferation.

biochemistry↗

bioRxiv2020

Sex-dependent pathways in hepatocarcinogenesis triggered by deregulated cholesterol synthesis

We uncover novel pathways of sex-dependent hepatocarcinogenesis due to chronic repression of cholesterol synthesis at the lanosterol 14-demethylase (CYP51) step. The response to metabolic insult determined by global liver transcriptome, qPCR and sterol metabolite analysis, together with blood parameters, revealed molecular signatures that differ between females and males. The data deduced from the mouse model are highly relevant for humans. The dampened hepatic metabolism presents a hallmark of carcinogenesis, particularly in ageing females, with increased plasma cholesterol and HDL, and a substantial negative enrichment of transcription factors from lipid metabolism, such as NR1B1, LXR, LRH1, and FXR. Importantly, the carcinogenic signalling pathways (ECM-receptor interaction and PI3K/Akt) are positively enriched, albeit with sex-dependent gene targets. The activated TGF-{beta}, mTOR, Wnt, and estrogen signalling worsen the phenotype, with NFATC1/2 being central to the female phenotype. This collectively leads to activated cell death and diminished basal metabolism. In conclusion, our data underline sex as an important biological variable of hepatocarcinogenesis. We uncover novel cholesterol-dependent transcription factors and signalling pathways as cancer markers in the ageing females. SignificanceChronic repression of the late part of cholesterol synthesis provokes hepatocarcinogenesis with sex-dependent modulation of signalling pathways and transcription factors. Aging females show specific metabolic signatures and a more aggrevated phenotype of metabolism-related HCC.

cancer biology↗