Risk-of-bias assessment of mixed studies combining in vitro, animal, and/or translational models to evaluate anti-infective drugs and vascular ageing.
| Ref # | Study type | Drug/Class | Model/Population | Tools used | Key bias domains (summary) | Overall risk | Ageing effects (biomarkers) |
|---|---|---|---|---|---|---|---|
| [42] | Mixed | Antibiotic (salinomycin) | Senescent cancer cells (mouse + in vitro) | SYRCLE + narrative | Randomization not detailed; selective reporting; high-dose in vitro arm | Some concerns | Induced immune destruction of senescent cancer cells; ↓ SA-β-gal; ↑ immune clearance |
| [52] | Mixed | Bactericidal antibiotics | Mammalian cells + animal validation | Narrative + SYRCLE | No blinding in animal arm; supratherapeutic dosing in vitro | High | ↑ Mitochondrial dysfunction, ↑ ROS, ↑ DNA damage—hallmarks of vascular ageing |
| [91] | Mixed | Chloroquine | Middle-aged mice + in vitro assays | SYRCLE + narrative | Attrition not fully reported; selective reporting in vitro | Some concerns | ↑ Autophagy modulation; ↓ proteasome activity; lifespan extension in mice |
| [32] | Mixed | Acyclovir | In vivo (mouse) (male C57BL/6N mice, HFD + STZ-induced diabetes)In vitro (HepG2) mechanistic | SYRCLE + narrative | Random allocation to groups reported; small sample size; blinding and allocation concealment not clearly described; outcomes (glucose, lipids, oxidative stress) fully reported; no obvious selective reporting | Some concerns/moderate | ↓ Fasting glucose, ↓ HbA1c, ↓ HOMA-IR; ↓ serum TG, T-CHO, LDL-C; ↑ hepatic T-SOD, ↓ MDA; ↑ glucose uptake in HepG2; ↓ ROS and AGEs; activation of PKM1 → AMPK/SIRT1 axis (metabolic/mitochondrial hallmarks relevant to vascular ageing) |
| [85] | Mixed | DHA (antimalarial/anti-infective) | In vivo (rat fibrosis) + in vitro (HSC culture) | Narrative + SYRCLE | Liver tissue (not vascular)• Fibrosis model, not natural aging• Drug regimen in pathological context, not typical antibiotic use• No vascular cells/vascular function measured | Moderate relevance (mechanistic hypothesis) | > 80% of p16+ cells were also p53+; < 9% Ki-67+ (implies senescence, not proliferation)- DHA dose-dependently reduced collagen deposition/fibrosis area vs. fibrotic controls (p-values significant, exact values in original figures)- In vitro, DHA increased G2/M-phase cells and decreased S-phase cells (quantitative shift vs. control)- Upregulation of senescence markers (p53, p16, p21, HMGA1) and increased SA-β-gal positive cells after DHA |
| [146] | Mixed | Nitazoxanide (antiprotozoal/antiparasitic) | Cell lines (senescence induced by bleomycin or D-galactose), and mice with bleomycin-induced pulmonary fibrosis | Narrative + SYRCLE | The model is lung, not vascular tissue• Fibrosis induced by chemical injury, not physiological aging• No data on vascular cells or vascular remodeling• Translational relevance to vascular aging inferred, not demonstrated | Moderate to high risk/indirect relevance | ↓ Senescence (fewer SA-β-gal + cells, preserved proliferation), ↓ SASP, preserved nuclear SIRT1, inhibited PI3K-WIPI1 pathway, attenuated fibrosis and collagen deposition in lungs |
Tools applied included SYRCLE (animal), narrative appraisal (in vitro), or a combination. Ageing effects included mitochondrial dysfunction, autophagy modulation, senescence clearance, ROS induction, and immune-mediated removal of senescent cells (n = 6). ↑: increased/upregulated/elevated, ↓: decreased/downregulated/reduced. AMPK: AMP-activated protein kinase; DHA: dihydroartemisinin; HSC: hepatic stellate cell; MDA: malondialdehyde; PKM1: pyruvate kinase M1; ROS: reactive oxygen species; SASP: senescence-associated secretory phenotype; T-SOD: total superoxide dismutase.
The authors thank Lusaka Apex Medical University, School of Health Sciences, and Faculty of Medicine, for their academic and institutional support towards the completion of this manuscript.
CWS: Conceptualization, Writing—original draft, Writing—review & editing. JM: Data curation, Resources, Writing—review & editing. SKD: Methodology, Writing—review & editing, Supervision. AC: Data curation, Visualization, Writing—review & editing. LS: Validation, Resources, Writing—review & editing. KM: Investigation, Formal analysis, Writing—review & editing. All authors read and approved the submitted version.
The authors declare that they have no conflicts of interest.
Not applicable.
Not applicable.
Not applicable.
The dataset used during the study is available from the corresponding author upon request.
No funding was received for this manuscript.
© The Author(s) 2026.
Open Exploration maintains a neutral stance on jurisdictional claims in published institutional affiliations and maps. All opinions expressed in this article are the personal views of the author(s) and do not represent the stance of the editorial team or the publisher.