Current Molecular Medicine - Online First

Obesity is a major risk factor for metabolic and cardiovascular disorders. Recently, emerging biomarkers, such as the Visceral Adiposity Index (VAI) and Lipid Accumulation Product (LAP), have garnered attention for their utility in assessing visceral obesity. Bilirubin, a potent endogenous antioxidant, has been associated with protective effects against various diseases. This study aims to investigate the relationship between serum total bilirubin (STB) levels and VAI/LAP in adults.

This cross-sectional study utilized data from the National Health and Nutrition Examination Survey (NHANES) collected between 2003 and 2020. The calculation of VAI and LAP was performed computationally. Weighted multivariate regression models were used to explore the potential correlation between STB levels and VAI or LAP. RCS curves were used to identify the potential non-linear relationship. Moreover, subgroup analyses were conducted to examine heterogeneity across different populations.

The analysis included a cohort of 10,625 individuals aged 20 to 85 years. Both unadjusted and adjusted statistical models revealed a significant negative association between STB levels and VAI or LAP (all P< 0.001). RCS indicates that these relationships are linear. Subgroup analyses identified particularly strong associations in non-smokers aged 20-59 without hypertension/diabetes (P < 0.05).

Our study's strengths include the use of nationally representative data with appropriate weighting, comprehensive adjustment for confounding variables, and pioneering research on the link between serum bilirubin levels and visceral fat indices, which may indicate early metabolic risk markers. This finding highlights the significant role of bilirubin in body fat distribution and lipid metabolism.

This study revealed that STB was associated with VAI or LAP among the specific general American population aged 20-59 without hypertension/diabetes. Further prospective investigations are warranted to clarify the temporal relationship between STB and novel obesity indices.

Pancreatic Ductal Adenocarcinoma (PDAC) is one of the most malignant gastrointestinal tumors. M1 macrophage, a subtype within the Tumor Microenvironment (TME), plays a vital role in the development of cancer. Despite its anti-tumoral functions, the specific mechanisms of its action remain incompletely understood.

The effect of M1 macrophages on the proliferation ability and cell viability of PDAC cells was evaluated by Cell Counting Kit-8 (CCK-8) cell proliferation assay, cell clone formation assay, and flow cytometry. Western blot, qRT-PCR, confocal microscope, RNA-sequencing, and transmission electron microscope were performed to assess lipid peroxidation and ferroptosis level of PDAC cells in the context of M1 macrophage or TNF-α.

M1 macrophages inhibited cell proliferation and promoted cell death of PDAC cells, in which ferroptosis played a vital role. Mechanistically, Tumor Necrosis Factor-alpha (TNF-α) released by M1 macrophages binds to the TNFR1 receptor on pancreatic cancer cells, activating the p38 MAPK signaling, which upregulates Acyl-CoA Synthetase Long-chain family member 4 (ACSL4) expression, a critical lipid metabolism enzyme linked to ferroptosis, thereby promoting ferroptosis. Knockdown of ACSL4 or TNFR1 significantly reduced TNF-α-induced ferroptosis.

TNF-α is a major inflammatory cytokine and is mainly generated by macrophages and T lymphocytes. It is involved in many pathological processes, such as inflammatory diseases, autoimmune diseases, and cancer. Studies have shown that the administration of recombinant TNF-α can induce tumor regression in mice with sarcomas. In our study, systemic injection of TNF-α slowed the tumor growth in nude mice, but with no significant difference compared with the control group, which may partially be attributed to its angiogenic activity. TNF-α signals via two distinct membrane-binding receptors, TNFR1 and TNFR2, which regulate various diseases. In pancreatic cancer, the role of TNF-α is complex and poorly understood. In a previous study, they found that exogenous systemic administration of human TNF-α, which interacted with murine TNFR1, significantly increased overall tumor growth in the Panc02-PDAC model. Intriguingly, the loss of TNFR1 led to an impediment of immune cell infiltration into the tumor and impaired immunosurveillance, which accelerated tumor growth. This suggests that TNFR1 exerts both pro-tumoral and anti-tumoral functions in the Panc02-PDAC model, but the overall outcome is likely dependent on the spatiotemporal availability of TNF-α. However, systemic TNF-α injection can lead to severe side effects in animals, limiting its further application. In a recent study, TNFR2 was found to promote tumorigenesis and progression in the KPC-PDAC model. Knockdown of TNFR2 or pretreatment with an anti-TNFR2 antibody could significantly slow the tumor progression and incidence. In our study, TNFR2 was found to have a low expression in pancreatic cancer cells and was barely detected with the failure of knockdown. However, the cell lines used in the former study were established from a KPC mouse model, while our experiments were conducted using human PDAC cell lines. Contrary findings are possible as cell lines originate from two different species. However, we will further investigate the mechanism of this difference.

In summary, this study revealed that M1 macrophages could induce ferroptosis in pancreatic cancer cells through secreting TNF-α, indicating a potential therapeutic option for PDAC.

Lung cancer remains the leading cause of cancer-related mortality. Determining the T790M resistance variants and epidermal growth factor receptor (EGFR) mutations is crucial for personalized treatment, especially when using targeted therapies.

This review article aims to comprehensively compare some of the various diagnostic techniques associated with liquid biopsies, such as cell-free DNA (cfDNA) for T790M and EGFR mutant identification. It also aims to evaluate their pertinence in clinical settings, as well as their sensitivity and specificity to determine how effectively they monitor treatment response and resistance.

A literature search was conducted using databases including PubMed, Scopus, and Web of Science. The keyword list included “EGFR mutations,” “T790M resistance,” “liquid biopsy,” “COLD PCR,” “NGS,” “ddPCR,” “BEAMing,” and other methods. The effect of these studies on diagnostic technologies for identifying EGFR mutations was assessed in terms of clinical practice, methodological accuracy, and significance. Sensitivity, specificity, clinical applicability, cost analysis, turnaround times, and ease of integration into clinical workflows were used as parameters for evaluation based on the literature.

There are advantages and disadvantages to cfDNA monitoring strategies for treatment response and resistance, as well as to the assessment of sensitivity, specificity, and clinical applicability for identifying EGFR mutations.

Advanced techniques such as COLD-PCR, LC-MS, qPCR, NGS sequencing, Sanger sequencing, PNA microarrays, the Allele-Specific Competitive Extension (ASCE) real-time PCR assay, and nanopore technology are necessary for personalized lung cancer management. However, depending on the objective of the work, the suitable method should be selected based on its benefits and drawbacks.

Colon cancer is a highly prevalent tumor with a high mortality rate worldwide. SLC41A2 is a member of the solute carrier family, but its role in colon cancer is still unclear.

The relationship between the expression level of SLC41A2 and clinicopathological features in colon cancer was investigated using data from the TCGA database. The differential expression genes of SLC41A2 were identified the potential role of SLC41A2 in colon cancer was analysed through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. By transfecting plasmids or siRNA to overexpress or knock down SLC41A2 in colon cancer cells, the effects of SLC41A2 on colon cancer cell proliferation, migration, and apoptosis were detected through EdU, MTT, wound-healing, Transwell, and JC-1 experiments. Western blot and ubiquitination experiments validated the regulation of GSK3β stability by SLC41A2. Rescue experiments and CCK8 assays confirmed the regulatory effect of SLC41A2 on GSK3β.

Compared to normal tissues, SLC41A2 exhibited a lower expression level in colon cancer, and the expression levels of SLC41A2 were correlated with the stage and Tumor Node Metastasis (TNM) classification. GO and KEGG analyses displayed that SLC41A2 primarily affected the growth factor activity and Wnt signaling pathway. Furthermore, elevated expression of SLC41A2 notably decreased the proliferation, migration and invasion of colon cancer cells, along with increased apoptosis. The overexpression of SLC41A2 and rescue experiments confirmed that SLC41A2 enhances the protein stability of GSK3β by inhibiting its ubiquitin-proteasome degradation and causes the upregulation of GSK3β, thereby suppressing the progression of colon cancer.

SLC41A2 was lowly expressed in colon cancer tissues or cells. By inhibiting the ubiquitin-proteasome degradation of GSK3β, SLC41A2 can significantly upregulate the expression of GSK3β, which ultimately suppresses the proliferation and migration of colon cancer cells.

Studies have shown that abnormal stress is a significant inducer of Intervertebral Disc Degeneration (IVDD). Although traction force is commonly used to delay IVDD, its effects on Nucleus Pulposus Cells (NPCs) and their secreted exosomes remain unclear. In addition, this study systematically revealed the relationship between miR-8485 and IVDD for the first time.

Cellular experiments were performed using a Flexcell cell stretching platform to apply traction force to NPCs. After optimizing loading parameters, NPC-derived exosomes (NPCs-exo) were isolated and subjected to miRNA high-throughput sequencing. Differentially expressed miRNAs were identified, and their regulatory effects on the Wnt/β-catenin pathway were investigated. Ex vivo rabbit spinal samples were used to validate the cellular experimental results under traction force loading.

NPCs-exo were found to be internalized by NPCs, and traction force promoted NPCs-exo secretion. High-throughput sequencing and differential expression analysis identified miR-8485 as a differentially expressed miRNA in NPCs-exo secreted under Cyclic Mechanical Tension (CMT) conditions. Dual-luciferase reporter assays confirmed the targeted regulatory relationship between miR-8485 and GSK-3β, as well as its involvement in the Wnt/β-catenin pathway-mediated regulation of NPCs degeneration. Ex vivo experiments, including morphological and immunofluorescence analyses, revealed that the traction group exhibited better morphology than the pressure group, with a more organized AF, NP, and higher NPCs content, though some loss persisted. Both groups showed significant differences in ECM markers (Collagen II, Aggrecan, MMP3) compared to the control (p < 0.05). Additionally, the traction group had significantly higher Collagen II and Aggrecan levels than the pressure group (p < 0.05).

CMT can promote the secretion of NPCs-exo, which are internalized by the NPCs. Through the delivery of miR-8485, NPCs-exo target and regulate GSK-3β, thereby enhancing Wnt/β-catenin pathway activity. This mechanism increases NPCs viability and extracellular matrix synthesis while suppressing apoptosis, ultimately delaying IVDD progression. Immunofluorescence staining in animal experiments confirmed that traction force effectively improves extracellular matrix expression in the IVD and mitigates stress-induced morphological alterations of the IVD.

The pathogenesis of diabetic kidney disease (DKD) is complex, and the specific biomarkers for detecting early diagnosis and monitoring kidney function deterioration are insufficient, which affects the prognosis of patients. The complement activation in glomeruli and renal interstitium contributes to the aggravation of DKD. Several key complement proteins, such as complement factor 3 (C3), CD59, and complement factor H-related protein 2 (CFHR2) were reported to be potential biomarkers for early diagnosis and prognosis for DKD.

In the current study, we focus on CFHR2, to investigate its capability and sensitivity as a DKD biomarker. As a non-invasive detection sample, urine has the characteristic of convenient sampling. In the current study, the urine samples were collected from three groups: diabetic patients without albuminuria, with micro-albuminuria, and macroalbuminuria, to analyze whether CFHR2 was associated with albuminuria concentration and declined renal function. Meanwhile, the urinary neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), and C3 were also examined by enzyme-linked immunosorbent assay (ELISA) to compare with CFHR2 to determine whether CFHR2 had an advantage in predicting the early detection and progression of DKD. The Spearman correlation analysis was performed for the correlation analysis. The receiver operating characteristic curve was used to analyze the diagnostic efficacy.

CFHR2 had superior diagnostic power to predict the early occurrence of DKD and disease progression, compared with NGAL, microalbumin, and C3 in urine.

CFHR2 has satisfactory potential to be a biomarker for early diagnosis and risk of progression of DKD.

The antiviral effects of type I interferons [IFNs] on respiratory syncytial virus [RSV]-infected airway epithelial cells have been identified. We aim to further reveal the mechanism of stat3 and kruppel-associated box-associated protein 1 [KAP1] in RSV-infected airway epithelial cells.

Using the A549 cell line, we investigated the impact of RSV infection, KAP1 overexpression, and stat3 inhibition with Stattic. Cell counting kit 8 assay was used to determine the viability, and enzyme-linked immunosorbent assay was applied to measure the levels of IL-6, IL-8, IL-1β, IFN-α, and IFN-β. Viral replication was tested via plaque assay. Meanwhile, quantitative real-time reverse transcription polymerase chain reaction or/and western blot were applied to measure the expressions of p-stat3 and KAP1 in the cells.

RSV infection repressed the viability, upregulated p-stat3 and KAP1 expressions, elevated levels of inflammation-related factors [IL-6, IL-8, IL-1β], and type I IFN immune response-associated factors [IFN-α, IFN-β], and promoted viral replication in A549 cells. Stattic attenuated the promoting effect of RSV on inflammation-related factors and viral replication, but enhanced its impact on IFN-α and IFN-β levels in the cells. More importantly, KAP1 overexpression reversed the effects of Stattic on viability, inflammation [IL-6, IL-8, IL-1β], type I IFN immune response [IFN-α, IFN-β], and viral replication in RSV-infected A549 cells.

Our findings unveil the pivotal role of stat3 inhibition in potentiating type I IFN-mediated antiviral responses against RSV in lung epithelial cells, revealing KAP1 as a potential therapeutic target for combating respiratory viral infections.