Biology, 17 April 2026
Research Background
The cornea is a transparent tissue that plays a crucial role in maintaining vision and protecting intraocular structures. Corneal damage caused by trauma, inflammation, or disease can lead to scarring, loss of transparency, and severe visual impairment. Although current treatments, such as eye drops and surgical interventions, have been widely applied, their regenerative efficacy remains limited and does not fully meet the need for complete restoration of corneal structure and function.
In this context, platelet-rich plasma (PRP), particularly umbilical cord-derived PRP (ucPRP), has emerged as a promising regenerative approach due to its high content of growth factors and low immunogenicity.
Materials and Methods
This study was designed to evaluate the effects of umbilical cord-derived platelet-rich plasma (ucPRP) on corneal epithelial wound healing through functional assays and molecular mechanism analysis.
Human corneal epithelial cells (hCEC) were cultured under standard conditions (37°C, 5% CO₂) in a specialized medium supplemented with growth factors. Cells from passages 3 to 7 were used to ensure biological consistency.
ucPRP was prepared from umbilical cord blood using a two-step centrifugation protocol to obtain platelet-rich plasma at an optimal concentration (~1×10⁹ platelets/mL). The final product was standardized, aliquoted, and cryopreserved for experimental use.
Wound healing capacity was assessed using a scratch assay model. Cells were grown to near confluence, and a linear scratch was created using a sterile pipette tip. Images were captured at 0 and 24 hours, and wound closure was quantified based on the area covered by migrating cells.
To investigate signaling mechanisms, experiments were conducted in the presence of specific inhibitors, including BAPTA-AM (intracellular Ca²⁺ chelator), PD98059 and U0126 (MEK/ERK inhibitors), SB203580 (p38 MAPK inhibitor), AG1478 (EGFR inhibitor), U73122 (PLC inhibitor), and suramin (purinergic receptor antagonist). These approaches enabled the identification of key signaling pathways involved in the healing process.
Intracellular Ca²⁺ dynamics were measured using the fluorescent probe Fluo-3/AM combined with confocal microscopy, allowing real-time monitoring of calcium signaling.
Protein expression, particularly aquaporins, was evaluated by Western blot for quantification and immunofluorescence for cellular localization.
Additionally, membrane water permeability was measured using stopped-flow light scattering, while H₂O₂ diffusion was assessed using the HyPer7 fluorescent biosensor in live cells.
Key Findings
The results demonstrated that ucPRP strongly promotes corneal regeneration through multiple mechanisms:
- Accelerated wound closure: ucPRP significantly enhances cell migration and proliferation, with optimal effects at a concentration of 10%.
- Activation of intracellular calcium signaling: ucPRP induces a rapid and transient increase in intracellular Ca²⁺, primarily due to extracellular calcium influx.
- Role of purinergic receptors: calcium signaling is highly dependent on purinergic receptor activation, which plays a central role in wound healing.
- Activation of the MAPK/ERK pathway: this pathway is a key regulator of cell proliferation and migration and is completely inhibited by specific blockers.
- Increased membrane permeability: ucPRP significantly enhances water permeability (~55%) and H₂O₂ diffusion across the cell membrane.
- Upregulation of Aquaporin-1 (AQP1): AQP1 expression increases by approximately 40% following ucPRP treatment, contributing to water transport, redox signaling, and cell motility.
Scientific Interpretation
This study elucidates the molecular mechanisms underlying ucPRP-mediated corneal regeneration, highlighting a multifactorial process involving:
- Activation of Ca²⁺ signaling via purinergic receptors
- Stimulation of the MAPK/ERK signaling cascade
- Upregulation of AQP1, enhancing water transport and redox signaling (H₂O₂)
Notably, AQP1 may function not only in water homeostasis but also as a “peroxiporin,” facilitating oxidative signaling essential for wound healing. However, the direct link between AQP1 and ERK activation requires further investigation.
Conclusion and Applications
ucPRP represents a promising biological therapy for corneal injuries, promoting regeneration through modulation of intracellular signaling and membrane transport functions.
These findings suggest potential applications of ucPRP in:
- Treatment of chronic corneal ulcers
- Ocular surface injuries
- Diseases associated with impaired corneal epithelial regeneration
Nevertheless, further in vivo studies and clinical trials are necessary to confirm its efficacy and safety before widespread clinical application.Top of FormBottom of Form
References
Martinotti, S., Pellavio, G., Bonsignore, G., Balbo, V., Mazzucco, L., Laforenza, U., & Ranzato, E. (2026). Umbilical Cord PRP Accelerates Corneal Wound Healing via AQP1 Upregulation and Calcium Signaling. Biology
Source: Biology
