Mycophenolate mofetil, one of the most widely prescribed immunosuppressants in organ transplantation, appears to interfere with blood vessel development in zebrafish embryos, according to a new research article published in Biological Research. The study, led by Mengqi Wan and colleagues with corresponding author Zigang Cao, found that exposure to the drug at concentrations of 10, 20, and 40 micrograms per liter produced a strikingly region-specific pattern of vascular disruption: excessive, irregular branching in the intersegmental vessels that run along the trunk, alongside inhibited and disorganized growth of the cerebrovasculature in the brain. The researchers traced this divergent phenotype to inhibition of two molecular targets, the enzyme inosine 5′-monophosphate dehydrogenase and the mitogen-activated protein kinase signaling pathway.
The findings matter because MMF, a prodrug of mycophenolic acid, has been considered relatively safe compared with other immunosuppressive agents. Approved by the US Food and Drug Administration in 1995 for preventing kidney transplant rejection, it is now used in regimens for various organ transplants and in autoimmune diseases such as lupus and myasthenia gravis. Its main advantages are the absence of liver and kidney toxicity and lower bone marrow toxicity than alternatives, though it carries known gastrointestinal side effects such as diarrhea and gastritis, an increased risk of cytomegalovirus infection, leukopenia, and mild anemia. According to the authors, the probability of toxic effects is higher in younger patients, which is why the drug is used more cautiously in that group. Yet few studies had previously assessed its effects on the developing vasculature.
To fill that gap, the team turned to zebrafish, a standard model in toxicology. Zebrafish embryos develop externally and are transparent, allowing vascular development to be monitored from the earliest stages, and the anatomy and molecular mechanisms of their vascular system closely resemble those of higher vertebrates. The researchers used transgenic lines, Tg(fli1a:EGFP) to visualize blood vessels and Tg(TP1:EGFP) to report Notch signaling activity, exposing embryos beginning at 24 hours post-fertilization for three consecutive days, with daily renewal of the drug solution. Each experiment was repeated at least three times.
The morphological consequences were clear. Compared with controls treated with the solvent dimethyl sulfoxide alone, MMF-exposed larvae showed significantly shorter body length, cerebral hemorrhage, pericardial edema, and delayed yolk sac absorption, with effects particularly evident in the 10 microgram per liter group. As drug concentration increased, body length decreased, yolk sac absorption was further delayed, and the pericardial area enlarged. Under laser confocal microscopy, the intersegmental vessels displayed irregular hyperbranching, with small branch vessels becoming more numerous as concentration rose. The cerebrovasculature showed the opposite trend: inhibition was apparent from the lowest dose onward, and cerebral vessel development became irregular and asymmetric, in sharp contrast to the symmetric patterning seen in controls.
The researchers then probed the molecular machinery behind these defects. Quantitative PCR revealed that expression of vascular endothelial growth factor genes, including VEGFaa, VEGFab, VEGFc, and VEGFd, and of receptor tyrosine kinase genes including Flt1, Flt4, Kdr, and Kdrl, was affected to varying degrees. Genes tied to the MAPK cascade, a pathway central to cell proliferation, differentiation, apoptosis, and angiogenesis, were downregulated across the board: MAPK1, MAPK3, MAPK8b, MAPK9, p38a, and p38b all showed reduced expression. MMF also significantly downregulated IMPDH2, the rate-limiting enzyme in the de novo synthesis of guanine nucleotides that the drug targets noncompetitively and reversibly to suppress lymphocyte proliferation.
Oxidative stress emerged as another facet of the response. Reactive oxygen species staining showed green fluorescence increasing with drug concentration, superoxide dismutase activity trended downward, and malondialdehyde levels rose, all indicating elevated oxidative stress. However, when the team attempted to rescue the vascular defects with the antioxidant astaxanthin, the treatment had no effect, and the larvae still showed irregular vessel proliferation. Similarly, activation of Notch signaling with sodium valproate slightly restored the fluorescent reporter signal in the Tg(TP1:EGFP) line, but failed to reverse the actual vascular abnormalities in the vessel-reporter line. Notch-related genes notch 1a, Her1, and Her6 were also downgraded in expression, yet the rescue failure indicated that MMF-induced angiogenesis defects are not primarily mediated through Notch signaling or oxidative stress.
The decisive results came from two other rescue experiments. When MMF-exposed embryos were co-treated with guanosine, the end product of the metabolic pathway that IMPDH inhibition blocks, the excessive intersegmental vessel branching was effectively normalized, confirming that the vascular dysplasia stems from guanine nucleotide depletion caused by IMPDH suppression. In a parallel experiment, co-treatment with diprovocim, an activator of the MAPK signaling pathway, effectively rescued the vascular disturbances compared with MMF treatment alone. Together, these results support the authors’ conclusion that MMF induces intersegmental vessel hyperbranching in zebrafish by inhibiting MAPK signaling and IMPDH, while VEGF and Notch pathways, though transcriptionally perturbed, do not drive the phenotype.
Perhaps the most intriguing question the study leaves open is why the same drug produces opposite vascular outcomes in different parts of the body: hyperbranching in the trunk and suppressed growth in the brain. The authors propose several hypotheses grounded in published literature. Differential basal expression of IMPDH isoforms between trunk and cerebral endothelial cells could make some vascular beds more sensitive to GTP depletion than others. Distinct MAPK isoform profiles across vascular beds might lead to opposite angiogenic outcomes when the pathway is inhibited. Tissue-specific thresholds in VEGF signaling could also contribute to the discrepant responses. The researchers explicitly state that the underlying mechanism remains a limitation of the study and warrants further in-depth investigation.
The work carries a note of caution for clinical practice rather than an immediate change in care. MMF is already known to be teratogenic, and fetuses born to pregnant women with a history of exposure during pregnancy have been shown to have various defects, which is why the drug carries strict pregnancy warnings. What this study adds is a mechanistic account, in an animal model, of how the drug disrupts embryonic vascular development through IMPDH-dependent guanine nucleotide synthesis and MAPK signaling, and it demonstrates that guanosine supplementation and MAPK activation can alleviate the resulting abnormalities in zebrafish. The authors say the findings provide a theoretical basis for the clinical use of MMF. As with all zebrafish toxicology work, the results are model-based observations at defined exposure concentrations and durations; translating them to human developmental risk will require further research, particularly given that the mechanistic explanation for the divergent trunk-versus-brain phenotype is still hypothetical.
By combining live imaging of transgenic embryos, transcript-level analysis of angiogenic pathways, oxidative stress assays, and pharmacological rescue experiments, the team built a converging body of evidence pointing to a specific metabolic and signaling vulnerability.
Subject of Research: Effects of the immunosuppressant mycophenolate mofetil on vascular development in zebrafish embryos
Article Title: Mycophenolate mofetil induces angiogenesis disorders in zebrafish embryos by inhibiting the mitogen-activated protein kinase signaling pathway
Article References: Wan, M., Xiao, Z., Shi, X., Ye, Z., Liu, J., Liu, X., Li, H., Liao, X., Zhang, S., Xiao, J., & Cao, Z. (2026). Mycophenolate mofetil induces angiogenesis disorders in zebrafish embryos by inhibiting the mitogen-activated protein kinase signaling pathway. Biological Research, 59(1), Article 57. https://doi.org/10.1186/s40659-026-00717-7
Image Credits: AI Generated
DOI: 10.1186/s40659-026-00717-7
Keywords: mycophenolate mofetil, zebrafish, angiogenesis, MAPK signaling, IMPDH, immunosuppressants, vascular development, oxidative stress, guanosine, teratogenicity, embryotoxicity, transplantation
News Source: Drew Townsend. (October 30, 2026). Common Immunosuppressant Disrupts Blood Vessel Growth in Zebrafish Embryos. Scienmag.
