With age, the skin undergoes progressive structural and functional decline [1]-[3]. The epidermis thins due to diminished keratinocyte proliferation, yielding slower stratum corneum renewal. Diminished lipid biosynthesis impairs the permeability barrier. Senescent dermal fibroblasts display reduced proliferation as well as lower synthesis of collagens and elastin, coupled with upregulated matrix metalloproteinase activity, which all precipitates extracellular matrix catabolism. Lower levels of hyaluronic acid and proteoglycans impair hydration, while reduced vascularisation limits nutrient delivery. Flattening of the dermal-epidermal junction further impairs nutrient and cellular exchange. Collectively, these changes result in visible ageing signs, including wrinkles, sagging, loss of elasticity, dryness, and uneven skin tone.

In an ever-ageing global population, visible hallmarks of cutaneous senescence constitute a mounting aesthetic and psychosocial burden [4]. The anti-ageing cosmetics sector is accordingly witnessing exponential growth [5]. Given the pivotal role of oxidative stress in senescence, antioxidants constitute core actives in many formulations [6][7]. Beyond this classical approach, retinol—the alcohol derivative of vitamin A—has been regarded as the reference compound for topical rejuvenation [8]. Being liposoluble, retinol penetrates the stratum corneum to reach the viable epidermis, albeit with limited efficiency. By stimulating keratinocyte proliferation, differentiation, and keratinisation, it enhances epidermal thickness, strengthens the stratum corneum, and improves barrier function [9][10]. Within the dermis, retinol promotes extracellular matrix synthesis, particularly collagen production, thereby increasing elasticity and reducing fine wrinkles [9][10]. Collectively, these actions account for its ability to reverse visible ageing signs [9][11].

Although a natural compound, the widespread use of retinol has revealed that its topical application can elicit erythema, xerosis, scaling, and desquamation [12][13]. Similar adverse reactions occur with synthetic retinoids, underscoring their limited tolerability. These drawbacks have prompted the search for better-tolerated alternatives. Bakuchiol—a meroterpene phenol derived from Psoraleacorylifolia—has emerged as a prominent plant-based retinol analogue, mimicking retinoid-like gene expression and clinical outcomes in photodamaged skin [14][15]. However, bakuchiol is not entirely devoid of adverse effects [16][17]. In this context, a Harungana madagascariensis extract (HME) has garnered interest.

In human fibroblasts, HME induces a broad transcriptional response that largely overlaps with that induced by retinol and includes upregulation of retinol receptors [18]. When formulated in a cream and topically applied to skin explants artificially aged by UV irradiation, it leads to higher levels of total collagen and type I collagen, an effect not observed with a retinol-containing cream. These findings were confirmed by the clinical evaluation of an HME-containing cream, which demonstrated improvements in multiple ageing parameters, including wrinkle depth, firmness, elasticity, and skin tone homogeneity, largely mimicking results obtained with retinol [19]. Therefore, HME appears a promising functional analogue of retinol, with potent beneficial effects on the dermis and an excellent tolerability profile, as no adverse events have been reported to date.

Reinforcing the dermis is crucial given the central role of the extracellular matrix degradation in the onset and severity of ageing signs. Indeed, it is its progressive breakdown diminishes skin strength and elasticity, ultimately resulting in the formation of static wrinkles and dynamic folds [20]. Nevertheless, a comprehensive anti-ageing approach should also strengthen the skin barrier function. During screenings for natural compounds reinforcing the epidermis, we identified a sea buckthorn (Hippophae rhamnoides L.) seed extract (BSE). We herein characterise the immunohistochemical effects of HME and BSE on artificially aged skin explants, alongside assessment of their synergistic action when combined.

Results demonstrate that HME increased dermal COL-I and CRABP-II marker levels, also enhancing epidermal CRABP-II and Ki67 levels. BSE, the effects of which are characterised here for the first time, upregulated dermal CRABP-II (+43%) and COL-I (+28% - 56%), whilst independently enhancing epidermal CRABP-II (+52% - 58%) and Ki67 proliferation (+59% - 84%). Most strikingly, the HME-BSE combinations elicited superior quantitative responses versus individual extracts for both STRA6 (+39% - 49% vs. no effect for HME/BSE) and TJP1 (+91% - 140% vs. +72% HME; +27% BSE 0.1%).

The effects of HME were anticipated as it was previously reported to induce a retinol-like transcriptomic profile in fibroblasts and to increase the retinol receptor CRABP-II and COL-I proteins in fibroblasts and UV-aged skin explants, also leading to higher Ki67 levels in UV-aged skin explants [18]. Our findings further support that HME recapitulates retinol’s established effects: increasing dermal COL-I and CRABP-II [20] and enhancing epidermal CRABP-II and Ki67 levels [20][21].

BSE induced effects very similar to those of HME. Both positively impact CRABP-II levels in the epidermis and the dermis, also inducing an increase in dermal COL-I, showing a strong dermal impact, while also increasing epidermal Ki67. These effects are also similar to those observed with the retinol cream, although no dermal CRABP-II increase was evidenced under our conditions. It is thus tempting to propose that BSE represents another plant extract with retinol-like properties and that both HME and BSE independently stimulate retinol effectors. Nevertheless, while BSE and HME were formulated in the same vehicle and are directly comparable, the retinol cream has a different vehicle of undisclosed composition and probably contains other active ingredients besides retinol. Comparisons should thus be made cautiously, although the results obtained align with the known impact of retinol [20][21]. Besides, only a few proteins were evaluated, and drawing firm conclusions about BSE’s retinol-like properties will require more thorough studies.

Nevertheless, the suggestion that both extracts possess retinol-like properties is further supported by their combined effects on CRABP-II. This combination did not increase epidermal CRABP-II levels, whereas each extract alone did. In the dermis, only the combination with the highest BSE concentration had an effect, whereas HME and all concentrations of BSE alone were effective. Besides suggesting potent effects of BSE on the dermis, these results suggest some interference between the extracts when combined. Whether a competitive interaction is involved remains to be elucidated, but these findings suggest that both extracts might interfere with the same CRABP-II target.

Interestingly, results also highlighted a response for STRA6 only when both extracts were combined. Besides, the expression level of TJP1 with the combination of extracts is superior to that achieved with each extract alone. These results suggest additive, and possibly synergistic, epidermal effects. Although multi-donor consistency lends plausibility to such an additive/synergistic effect, differences across conditions were limited, restricting analyses to pairwise comparisons against UV-irradiated untreated controls and preventing multiple comparative analyses that would have strengthened this conclusion.

The additive/synergistic effect suggests that the extract combination coordinately contributes to epidermal reinforcement via a dual mechanism. On one hand, STRA6 is a membrane transporter and a cell-surface receptor that selectively binds all-trans-retinoic acid, constituting a central regulator of intracytoplasmic retinoid levels. When present in larger amounts, it enables higher intracellular retinoid concentrations, ultimately resulting in keratinocyte proliferation and differentiation [22][23]. STRA6 upregulation should thus strengthen the epidermis by favouring its development. On the other hand, TJP1 is a scaffolding protein and component of the tight junction complex. This complex creates a dynamic mechanical barrier in the stratum granulosum, which—among other functions—regulates permeability to ions and solutes, also contributing to resistance against mechanical and chemical stressors [24][25]. Increased TJP1 levels should, therefore, strengthen barrier function by stabilising tight junction complexes, reducing transepidermal water loss (TEWL), limiting solute permeability, and enhancing cellular cohesion. Both extracts also individually upregulated Ki67, a marker of basal epidermal cell proliferation [26]. Although not showing any additive/synergistic effect on this latter marker, the stimulation observed with each extract should further contribute to a strong reinforcement of the epidermis, supporting the notion that the epidermis is an important site of action of HME and BSE.

The additive/synergistic effect might appear to contradict the negative interaction we highlighted for HME-BSE combination on CRABP-II. Indeed, if such a competitive interaction exists between the extracts, one might expect reduced retinol-like properties when extracts are combined. This was not the case, as the extract combination induced changes within the range of those induced by each extract alone. Alternative explanations must therefore be sought, such as a CRABP-II-independent mode of action. This seems plausible, as Harungana madagascariensis and Hippophae rhamnoides seeds are both known to be rich in secondary metabolites, including flavonoids, proanthocyanidins, xanthones, and tocopherols [27][28]. Such pleiotropic bioactivities might explain the preserved retinol-like efficacy despite CRABP-II antagonism, with synergy emerging via complementary modulation of STRA6/TJP1.

Taken together, this study suffers limitations that will require further work to confirm or refute our conclusions. Among these limitations are the modest effect sizes observed, highlighting the substantial inter-individual variability of skin explant responses to the various treatments, including responses to the retinol cream. This prevented detection of significant differences across treatments whilst the multi-donor design represents a methodological strength. Moreover, the analysis of only six markers limits the mechanistic breadth of the study and precludes firm claims regarding BSE’s retinol-like properties. Furthermore, the use of explants from only six Caucasian female donors aged 26 - 47 years limits the generalizability of the present findings to older individuals and to individuals with more diverse ethnic backgrounds. Although standardized UVA/UVB irradiation was used to induce a controlled photoageing-like state, this acute ex vivo UV challenge does not fully reproduce the complex intrinsic and cumulative extrinsic ageing processes occurring in older skin. Future work should encompass broader proteomic/transcriptomic profiling alongside clinical trials to validate bonafide retinol-like efficacy of BSE. Parallel efforts to identify BSE’s active compounds would also be of interest to dissect synergy, warranting both invitro mechanistic and invivo translational studies.

This study provides the first evidence that Hippophae rhamnoides seed extract (BSE) upregulates key retinol-responsive markers (CRABP-II, COL-I, Ki67), mirroring Harungana madagascariensis extract (HME) effects and suggesting retinol-like anti-ageing potential. Considering the irritancy risks of retinol and its derivatives, there is a growing demand for gentler alternatives that also align with consumer preference for natural actives. Remarkably, HME-BSE synergy elevates epidermal STRA6/TJP1, complementing HME’s dermal dominance with BSE’s epidermal contributions to yield superior barrier/proliferative reinforcement, positioning the HME-BSE combination as a promising dual-compartment anti-ageing strategy.

The authors would like to thank Dr. Philippe Crouzet, Estium-Concept, for providing scientific writing services.

SB, MCB and NS conceived and designed the study under supervision of RF. LB contributed to project administration. MCB and LC performed experiments, collected and analyzed the data, and conducted statistical analysis. SB, MCB and RF contributed to the interpretation of the results. SB, MCB and RF wrote the initial draft of the manuscript. All authors read and approved the final version of the manuscript.