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Article

Bioconverted Blueberry Extract Potentiates the Angiogenic and Endothelial Functions in Human Dermal Microvascular Endothelial Cells Under Oxidative Stress

by
Jung Un Shin
1,
Yun Hoo Jo
1,
Soo Ah Jeong
2,3,
Yeong Hwan Jeong
2,
Myeong Gwan Son
2,3,
Yoo Jeong Jeong
2,
Beong Ou Lim
2,3 and
Dong Wook Shin
1,*
1
Research Institute for Biomedical and Health Science, Kunkuk University, Chungju 27478, Republic of Korea
2
Department of Applied Biochemistry, Kunkuk University, Chungju 27478, Republic of Korea
3
Human Bioscience Corporate R&D Center, Human Bioscience Corp., Chungju 27478, Republic of Korea
*
Author to whom correspondence should be addressed.
Curr. Issues Mol. Biol. 2026, 48(2), 224; https://doi.org/10.3390/cimb48020224
Submission received: 31 December 2025 / Revised: 8 February 2026 / Accepted: 12 February 2026 / Published: 19 February 2026
(This article belongs to the Special Issue Molecular Insights into Food-Derived Natural Products and Their Biological Activities—2nd Edition)
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Abstract

Endothelial dysfunction induced by oxidative stress is a critical contributor to impaired microvascular homeostasis and skin aging. Blueberries are rich in polyphenolic compounds with antioxidant properties. However, whether bioconversion enhances their protective effects on endothelial function remains insufficiently explored. In this study, we investigated the beneficial effects of bioconverted blueberry extract (BBS) on human dermal microvascular endothelial cells (HDMECs). HDMECs were exposed to hydrogen peroxide (H2O2) to induce oxidative stress and subsequently treated with BBS. BBS significantly reduced H2O2-induced ROS accumulation and preserved mitochondrial membrane potential. Consistently, BBS markedly enhanced endothelial migration and tube-forming ability under oxidative stress conditions. Furthermore, BBS treatment significantly suppressed the overactivation of MAPK signaling pathways. Collectively, BBS effectively mitigated oxidative stress-induced endothelial dysfunction by restoring redox balance, preserving mitochondrial integrity, and promoting angiogenic function. Taken together, these findings suggest that bioconverted blueberry extract can be utilized as a functional ingredient for skin health and anti-aging.

1. Introduction

The integumentary and vascular systems are intimately interconnected, with the skin serving not only as a barrier against external insults but also as a highly vascularized organ that relies on microcirculatory support for homeostasis, repair, and regeneration [1,2]. The cutaneous microvasculature delivers oxygen and nutrients while removing metabolic waste, and its functional integrity is critical for maintaining skin physiology across the lifespan [3]. Dysregulation of vascular dynamics has been implicated in a variety of skin conditions, including impaired wound healing, chronic inflammatory dermatoses, and age-associated degeneration [4]. Moreover, accumulating evidence suggests that declines in microvascular function contribute to compromised epidermal barrier integrity, reduced skin perfusion, and an increased propensity for oxidative damage, collectively accelerating skin aging [5,6,7].
Endothelial cells lining the microvasculature play a central role in orchestrating these processes by acting as sensors and regulators of blood flow, inflammation, and angiogenic signaling [8,9,10]. Human dermal microvascular endothelial cells (HDMECs) have been widely utilized to investigate the mechanisms underlying angiogenesis, endothelial barrier maintenance, and interactions with perivascular and immune cells under both homeostatic and pathological conditions [11,12,13]. Given the pivotal role of endothelial metabolism in vascular health, exploring metabolic modulators that enhance endothelial resilience has become a research priority [14].
Bioconversion technologies have recently gained traction as an innovative approach to enhance the bioactivity and bioavailability of plant-derived compounds [15,16]. Bioconversion employs specific microorganisms or enzymatic processes to transform complex phytochemicals into more readily absorbable and functionally potent metabolites [17]. This strategy has been applied across food, nutraceutical, and biomedical research to generate functional metabolites from polyphenol-rich substrates such as grape, green tea, and berries [18,19,20]. Among these substrates, blueberries (Vaccinium corymbosum) are of particular interest due to their high content of anthocyanins and other phenolic compounds [21].
Previously, we successfully developed a propionic acid-rich bioconverted blueberry extract (BBS) and confirmed its biological efficacy [22]. In the present study, we extended our investigation to evaluate the effects of BBS on HDMEC function, in comparison with non-bioconverted blueberry (BNB), and using ascorbic acid (AA) as a positive control [23], with a focus on endothelial migration, resilience against oxidative stress, and angiogenic function and signaling. By leveraging an in vitro HDMEC model, we sought to elucidate whether bioconversion-enhanced blueberry metabolites could serve as functional modulators of dermal microvascular health. These findings provide mechanistic insight into their potential application for skin health and anti-aging strategies.

2. Materials and Methods

2.1. Blueberry Fermentation

A blueberry-based mixture (BNB: non-bioconverted blueberry) was prepared by combining blueberry juice, human bio science broth (HBS001; Mediogen, Jecheon, Chungcheongbuk-do, Republic of Korea), and distilled water in a ratio of 50:7:500. The mixture was heat-treated at 90 °C for 10 min, cooled to room temperature, and subsequently exposed to ultraviolet irradiation for 30 min in a vertical laminar flow cabinet (JSCB-900SL; JS Research, Gongju, Republic of Korea). The sterilized BB base mixture was diluted 10- to 20-fold with distilled water, and the pH was adjusted to 6.0 using sodium hydroxide (NaOH) and hydrochloric acid (HCl). The pH was continuously monitored throughout the fermentation process and readjusted as necessary to maintain a stable fermentation environment. Dietary fiber and minerals containing HBS were added to the diluted mixture, followed by inoculation with Lactiplantibacillus plantarum HBS01 at a concentration of 5–10% (v/v). Fermentation was conducted in a shaking incubator at 37 °C and 80 rpm for 24 h. Following fermentation, the mixture was subjected to low-temperature pasteurization at 65 °C for 1 h. The final fermentation product, blueberry supernatant (BBS: Blueberry Bioconversion short-chain fatty acids (SCFAs), was supplied by Human Bioscience (Chungju, Republic of Korea) and utilized for subsequent analyses.

2.2. Cell Culture

HDMECs (PromoCell, Heidelberg, Germany) were cultured in endothelial cell growth medium (PromoCell, Heidelberg, Germany) supplemented with 1% penicillin–streptomycin (Welgene Inc., Gyeongsan, Republic of Korea) at 37 °C in a humidified incubator with 5% CO2.

2.3. Cell Viability Assay

HDMECs were seeded into 96-well plates at a density of 1 × 104 cells per well and cultured for 24 h. The cells were then treated with BBS at various concentrations (0, 0.1, 0.5, 1, 2.5, and 5%) for an additional 24 h at 37 °C in a humidified incubator with 5% CO2. After treatment, the BBS-containing medium was removed, and the cells were washed twice with DPBS. Next, MTT labeling reagent (0.5 mg/mL in culture medium) was added to each well and incubated for 4 h at 37 °C under 5% CO2. Following incubation, 100 μL of dimethyl sulfoxide (DMSO) was added to each well to dissolve the formazan crystals, and the plates were incubated for 30 min at 5% CO2 incubator. Absorbance was then measured at 570 nm.

2.4. Wound Healing Assay

HDMECs were seeded into 6-well culture plates and cultured for 24 h. A linear scratch wound was generated in the center of each well using a sterile 200 μL pipette tip. The cells were then treated with 200 μM H2O2, 200 μM ascorbic acid (AA), BNB, or BBS at concentrations of 2.5%. After treatment, the cells were incubated for an additional 24 h under the same culture conditions. Phase-contrast images were captured at 0 and 24 h after treatment using a Nikon light microscope (Tokyo, Japan) to assess wound closure.

2.5. DCF-DA ROS Assay

Intracellular ROS production was determined using a Cellular ROS Assay Kit (Abcam, Cambridge, UK). HDMECs were plated in confocal dishes and allowed to attach for 24 h. The cells were then exposed to 200 μM AA, 2.5% BBS, or 2.5% BNB for 22 h, followed by exposure to 200 μM H2O2 for 2 h. After treatment, the cells were incubated with 10 μM 2′,7′-dichlorofluorescin diacetate (DCF-DA) for 20 min under light-protected conditions. The staining solution was subsequently removed, and the cells were washed with DPBS. Intracellular fluorescence was observed using a Nikon Eclipse Ti2 live-cell fluorescence microscope (Tokyo, Japan).

2.6. Measurement of Mitochondrial Membrane Potential

Mitochondrial membrane potential was evaluated using the JC-1 Mitochondrial Membrane Potential Assay Kit (Abcam, Cambridge, UK). HDMECs were plated in confocal dishes and maintained for 24 h. The cells were treated with 200 μM AA, 2.5% BBS, or 2.5% BNB for 22 h, followed by exposure to 200 μM H2O2 for 2 h. The culture medium was then removed, and the cells were incubated with 5 μM JC-1 dye for 30 min under light-protected conditions. After staining, the cells were washed with DPBS, and fluorescence images were captured using a Nikon Eclipse Ti2 live-cell fluorescence microscope (Tokyo, Japan).

2.7. Western Blot Analysis

HDMECs were seeded in 100 mm culture dishes and allowed to attach for 24 h. The cells were then treated with 200 μM H2O2, 200 μM AA, 2.5% BBS, or 2.5% BNB for 30 min. Following treatment, the cells were lysed in RIPA buffer and centrifuged at 12,000 rpm for 10 min at 4 °C. Protein concentrations were determined using a BCA protein assay kit (Thermo Fisher Scientific, Waltham, MA, USA). A total of 25 μg of protein from each sample was separated by SDS-polyacrylamide gel electrophoresis and transferred onto polyvinylidene difluoride membranes. The membranes were blocked with 5% skim milk in Tris-buffered saline containing 0.1% Tween-20 (TBS-T) for 2 h and then incubated overnight at 4 °C with primary antibodies against extracellular signal-related kinases (ERKs), p-ERK, p38, p-p38, stress-activated kinases c-Jun N-terminal kinase (JNK), p-JNK, and β-actin (Cell Signaling Technology, Beverly, MA, USA). After three washes with TBS-T, the membranes were incubated with horseradish peroxidase-conjugated secondary antibodies for 2 h, followed by three additional washes. Protein bands were detected using an enhanced chemiluminescence (ECL) reagent (Cytiva, Marlborough, MA, USA) and visualized with the Invitrogen iBright 1500 imaging system (Waltham, MA, USA). Densitometric analysis was performed using ImageJ software (version 1.53e, National Institutes of Health, Bethesda, MD, USA), and protein expression levels were normalized to β-actin.

2.8. Tube Formation Assay

Tube formation was assessed using an Angiogenesis Assay Kit (Cell Biolabs, San Diego, CA, USA). A pre-chilled 96-well plate was coated with 50 μL of Matrigel per well and allowed to polymerize for 30 min. HDMECs (2.5 × 104 cells per well) were resuspended in 150 μL of culture medium and seeded onto the Matrigel-coated wells. Tubular structures were observed after 6 and 24 h of incubation using a Nikon light microscope (Tokyo, Japan).

2.9. ELISA Assay

HDMECs were treated with 200 μM H2O2, 200 μM AA, 2.5% BBS, or 2.5% BNB and incubated for 6 h at 37 °C in a humidified atmosphere containing 5% CO2. After treatment, the cell culture supernatants were collected and used as samples for ELISA analysis. VEGFA secretion was quantified using a Human VEGFA ELISA Kit (Cat. NO. BMS277-2; Thermo Fisher Scientific, Waltham, MA, USA), according to the manufacturer’s instructions. Briefly, samples were added to microplates pre-coated with a capture antibody, followed by incubation with an enzyme-linked detection antibody to specifically detect VEGFA based on the sandwich ELISA principle. After substrate addition, the colorimetric reaction was measured at 450 nm using a microplate reader (BioTek, Winooski, VT, USA). VEGFA concentrations were determined by comparison with a standard curve.

2.10. Statistical Analysis

All results are expressed as the mean ± standard deviation (SD), with error bars representing SD from independent experiments. Statistical differences among groups were analyzed using one-way analysis of variance (ANOVA), followed by appropriate post hoc tests. All statistical analyses were performed using GraphPad Prism software (version 8.01; San Diego, CA, USA).

3. Results

3.1. Effects of BBS on HDMECs Viability and Proliferation

To assess the potential cytotoxicity of BBS in HDMECs, cell viability was measured using the MTT assay. HDMECs were exposed to various concentrations of BBS (0.1, 0.5, 1, 2.5, 5%) for 24 h. Cell viability remained above 90% at 5% concentration (Figure 1), indicating that BBS was not cytotoxic at this level. However, treatment with 5% BBS resulted in lower cell migration compared with treatment with 2.5% BBS. Therefore, a BBS concentration of 2.5% were chosen as a non-cytotoxic dose for the following experiments.

3.2. BBS Promoted Migration of H2O2-Damaged HDMECs

HDMECs were treated with 200 μM H2O2, 200 μM AA, 2.5% BBS, or 2.5% BNB. After 24 h, H2O2 treatment significantly delayed wound closure compared with the control. Notably, BBS treatment restored wound closure in H2O2-damaged HDMECs in a concentration-dependent manner, with 2.5% BBS showing the greatest effect (Figure 2A,B).

3.3. BBS Decreased Intracellular ROS Level in H2O2-Damaged HDMECs

Intracellular ROS levels in HDMECs were assessed using a DCF-DA assay, in which green fluorescence intensity is proportional to ROS accumulation. HDMECs were pre-treated with 200 μM AA, 2.5% BBS, or 2.5% BNB for 22 h, followed by exposure to 200 μM H2O2 for 2 h. As expected, a marked increase in green fluorescence was observed in the H2O2-damaged group, whereas BBS treatment significantly reduced fluorescence intensity. These results demonstrate that BBS attenuates intracellular ROS accumulation in H2O2-damaged HDMECs (Figure 3A,B).

3.4. BBS Restored Mitochondrial Membrane Potential (ΔΨm) in H2O2-Damaged HDMECs

The JC-1 assay was used to evaluate the effect of BBS on ΔΨm in H2O2-damaged HDMECs. In this assay, red fluorescence represents polarized, functional mitochondria, whereas green fluorescence indicates depolarized, dysfunctional mitochondria. H2O2 exposure significantly increased green fluorescence intensity compared with the control group, indicating mitochondrial depolarization. In contrast, BBS treatment enhanced red fluorescence, suggesting that BBS restored ΔΨm in H2O2-damaged HDMECs. Notably, BBS tended to confer slightly greater protection than BNB against H2O2-induced mitochondrial membrane potential impairment (Figure 4A,B).

3.5. BBS Reduced MAPK Phosphorylation in H2O2-Damaged HDMECs

To assess the inhibitory effects of BBS on MAPK phosphorylation under H2O2 stimulation, we analyzed the phosphorylation levels of ERK, p38, and JNK. As expected, H2O2 treatment markedly increased the phosphorylation of all three MAPKs. In contrast, BBS treatment significantly suppressed MAPK activation, whereas BNB showed little ability to suppress MAPK signaling overactivation (Figure 5A,B).

3.6. BBS Improved Tube Formation in H2O2-Damaged HDMECs

In the present study, H2O2 treatment significantly reduced tube length, mesh formation, and node number compared with the untreated control, indicating a marked impairment of angiogenic function. To assess the protective effect of BBS, HDMECs were co-treated with H2O2 and BBS. Notably, BBS markedly restored tube-forming ability in H2O2-exposed HDMECs, as evidenced by significant increases in tube length and node number relative to the H2O2-treated group. In addition, BBS tended to enhance the tube-forming ability to a greater extent than BNB (Figure 6A,B).
VEGFA secretion was evaluated by ELISA to support the pro-angiogenic effects of BBS at the molecular level. H2O2 treatment significantly decreased VEGFA secretion compared with the untreated control, whereas BBS treatment markedly increased VEGFA levels in H2O2-damaged HDMECs. Notably, VEGFA secretion in the BBS-treated group was significantly higher than that observed in the BNB-treated group (Figure 6C).

4. Discussion

Excessive intracellular ROS is a major driver of endothelial dysfunction, particularly under pathological or aging-related conditions [24,25]. Elevated ROS levels interfere with cytoskeletal dynamics and directional migration, thereby delaying wound repair [26]. Hydrogen peroxide (H2O2) has been shown to induce aging-related changes in the skin and vasculature [27]. Consistent with previous studies, H2O2 treatment significantly impairs endothelial cell migration [28]. In this study, BBS effectively restored wound closure and improved endothelial migratory capacity compared to the H2O2- and BNB-treated groups. Together, these results indicate that BBS promotes wound healing in H2O2-damaged HDMECs, which was consistent with the observed effects on cell viability.
Excessive accumulation of reactive oxygen species (ROS) triggers endothelial cell damage by activating regulated cell death pathways, including apoptosis, autophagy, necroptosis, and ferroptosis [29,30]. This ROS overload disrupts vascular homeostasis, leading to impaired endothelial integrity and vascular dysfunction [31,32]. In the present study, BBS significantly reduced H2O2-induced ROS accumulation and concomitantly enhanced endothelial migration. This finding suggests that BBS may restore the redox balance, which contributes to improved cellular motility rather than nonspecific proliferative effects.
Mitochondria play a central role in meeting cellular energy demands during energy-intensive biological processes such as angiogenesis [33]. Mitochondrial dysfunction disrupts cellular energy metabolism and redox balance, leading to excessive ROS accumulation and impairment of vascular homeostasis, which in turn compromises skin integrity [34,35,36]. Endothelial cells, including HDMECs, are particularly vulnerable to mitochondrial dysfunction, as mitochondrial integrity is essential for their adaptation to oxidative environments [37]. Loss of mitochondrial membrane potential (ΔΨm) results in reduced ATP production and enhanced ROS generation, thereby exacerbating endothelial dysfunction [38]. The preservation of ΔΨm in BBS-treated HDMECs indicates that BBS supports mitochondrial resilience, which is likely critical for sustaining energy-dependent processes such as migration and network formation.
The mitogen-activated protein kinases (MAPK) signaling pathway is activated in response to ROS [39,40]. MAPKs comprise growth factor-regulated ERKs, p38, and JNK [41,42]. MAPK signaling pathways are key redox-sensitive regulators of endothelial responses [43]. While it has been suggested that transient MAPK activation supports physiological angiogenic signaling, sustained activation under oxidative stress is associated with endothelial dysfunction. The attenuation of MAPK activation observed in BBS- treated cells suggests that BBS suppresses stress-associated MAPK overactivation, thereby facilitating functional recovery without broadly inhibiting essential signaling pathways.
Consistent with these molecular effects, BBS significantly improved tube formation capacity in HDMECs under oxidative stress. Endothelial cells have the intrinsic capacity to differentiate and form capillary-like networks when cultured on Matrigel [44]. The tube formation assay is a widely employed in vitro method for quantitatively assessing angiogenic activity by measuring parameters such as total tube length and the number of nodes (Nb nodes) [45,46,47]. Rather than indicating excessive angiogenic stimulation in the BBS-treated group, this enhancement reflects the restoration of endothelial network-forming ability, which requires coordinated regulation of redox homeostasis, mitochondrial function, and kinase signaling [48]. Compared with BNB, BBS treatment resulted in a significantly greater enhancement of endothelial tube formation, particularly in total tube length. These findings suggest that BBS exerts a more pronounced effect than BNB on endothelial network formation, a key functional indicator associated with improved skin vascular flow. Excessive activation of JNK under oxidative stress conditions has been associated with endothelial dysfunction and impaired angiogenic signaling [49]. In this context, the attenuation of JNK activation by BBS, likely attributable to bioconversion-induced compositional changes, may contribute to the restoration of VEGFA secretion and angiogenic capacity in H2O2-damaged HDMECs.
In our earlier work, high-performance liquid chromatography analysis confirmed that this bioconversion process enriches the fermented blueberry extract with short-chain fatty acids (SCFAs), with propionic acid identified as a predominant component at a concentration of 1.86 mg/mL [22]. SCFAs such as acetate, propionate, and butyrate have been reported to exert beneficial effects on endothelial cells, modulating intracellular redox balance, mitochondrial function, and stress-responsive signaling pathways [50,51,52]. Thus, we assume that SCFAs in BBS likely contribute to alleviating oxidative stress-induced dysfunction in HDMECs. In particular, SCFAs have been shown to influence mitochondrial metabolism and antioxidant capacity, which may be underlying the preservation of mitochondrial membrane potential and the suppression of stress-associated MAPK overactivation observed in BBS-treated cells [53].
Beyond SCFAs, microbial fermentation is known to promote the deglycosylation of blueberry anthocyanins, leading to the formation of more bioavailable aglycones and low-molecular-weight phenolic acids, such as gallic acid or protocatechuic acid [54]. Although these fermentation-derived metabolites were not directly quantified in the current study, their generation during microbial bioconversion has been well-documented and may further contribute to the enhanced bioactivity of BBS. We therefore propose that these BBS-specific metabolites act synergistically with propionic acid to confer superior antioxidant defense and mitochondrial protection compared with the original non-bioconverted blueberry extract (BNB), which predominantly contains less readily absorbable glycoside forms. Detailed comparative chemical and metabolomic profiling of BBS and BNB is currently prioritized for future studies to identify the specific mediators responsible for these synergistic effects.
Intracellular ROS levels were appropriately evaluated using the DCF-DA assay. Although this approach effectively measures overall ROS accumulation, the assessment of specific oxidative damage markers, such as 8-hydroxy-2′-deoxyguanosine for DNA oxidation or 4-hydroxynonenal for lipid peroxidation, would further strengthen the mechanistic association between oxidative stress reduction and endothelial functional recovery. Taken together, these considerations highlight the need for further studies to directly elucidate the molecular mediators underlying the endothelial protective effects of blueberry bioconversion-derived materials, particularly the role of SCFAs. Future investigations should aim to characterize the SCFA composition of BBS in detail and to evaluate the direct effects of individual SCFAs on HDMEC function under oxidative stress conditions. Such studies will provide a more definitive understanding of how SCFAs contribute to the restoration of endothelial migration and network-forming capacity observed in this study.
Collectively, these findings indicate that BBS restores endothelial functional competence under oxidative challenge by re-establishing redox balance, preserving mitochondrial integrity, and modulating stress-responsive MAPK signaling (Figure 7). Given the central role of oxidative stress-induced endothelial dysfunction in impaired wound healing and microvascular degeneration, BBS may serve as a promising functional modulator of dermal microvascular health.

5. Conclusions

In summary, BBS mitigated H2O2-induced oxidative stress and functional impairment in human dermal microvascular endothelial cells. These findings suggest that BBS serves as a functional modulator of dermal microvascular endothelial function under oxidative conditions.

Author Contributions

Conceptualization, J.U.S. and D.W.S.; methodology, J.U.S.; formal analysis, J.U.S.; investigation, J.U.S., Y.H.J. (Yun Hoo Jo), S.A.J., Y.H.J. (Yeong Hwan Jeong), M.G.S., and Y.J.J.; writing—original draft preparation, J.U.S.; writing—review and editing, B.O.L. and D.W.S.; supervision, D.W.S.; project administration, D.W.S.; funding acquisition, D.W.S. All authors have read and agreed to the published version of the manuscript.

Funding

This research was supported by the Regional Innovation System & Education (RISE) program through the Chungbuk Regional Innovation System & Education Center, funded by the Ministry of Education (MOE) and the Chungcheongbuk-do, Republic of Korea (2025-RISE-11-003-03).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in this study are included in the article. Further inquiries can be directed to the corresponding author.

Acknowledgments

The scientific illustration was created using BioRender (Toronto, ON, Canada).

Conflicts of Interest

Ms. Soo Ah Jeong, Mr. Myeong Gwan Son, and Professor Dr. Beong Ou Lim were employed by the company Human Bioscience Corporate R&D Center, Human Bioscience Corp. However, the authors confirm that there are no conflicts of interest to declare, and the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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Figure 1. Effects of BBS on the viability of HDMECs. Cell viability of BBS at various concentrations was assessed using the MTT assay. Cell viability was expressed as the percentage (%) relative to the untreated control group. Data are presented as mean ± SD (n = 3), with statistical significance denoted as # p < 0.05 relative to the control group.
Figure 1. Effects of BBS on the viability of HDMECs. Cell viability of BBS at various concentrations was assessed using the MTT assay. Cell viability was expressed as the percentage (%) relative to the untreated control group. Data are presented as mean ± SD (n = 3), with statistical significance denoted as # p < 0.05 relative to the control group.
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Figure 2. Effects of BBs on wound healing in H2O2-stimulated HDMECs. Cells were treated with 200 μM AA, 2.5% BBS, or 2.5% BNB in the presence of 200 μM H2O2 for 24 h. (A) Representative phase-contrast images showing wound closure in HDMECs at 0 h and 24 h (scale bar 50 μm). The blue lines indicated the scratch area. Images represent results from three independent experiments. (B) Quantitative analysis of the wound-closure area was performed using ImageJ software (version 1.53e). ‘+’ and ‘-’ indicate the presence and absence of the indicated substance, respectively. Data are presented as mean ± SD (n = 3), with statistical significance denoted as * p < 0.05 relative to the H2O2-treated group. ## p < 0.01 compared with the control group. No significant difference was observed between the BBS- and BNB-treated groups.
Figure 2. Effects of BBs on wound healing in H2O2-stimulated HDMECs. Cells were treated with 200 μM AA, 2.5% BBS, or 2.5% BNB in the presence of 200 μM H2O2 for 24 h. (A) Representative phase-contrast images showing wound closure in HDMECs at 0 h and 24 h (scale bar 50 μm). The blue lines indicated the scratch area. Images represent results from three independent experiments. (B) Quantitative analysis of the wound-closure area was performed using ImageJ software (version 1.53e). ‘+’ and ‘-’ indicate the presence and absence of the indicated substance, respectively. Data are presented as mean ± SD (n = 3), with statistical significance denoted as * p < 0.05 relative to the H2O2-treated group. ## p < 0.01 compared with the control group. No significant difference was observed between the BBS- and BNB-treated groups.
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Figure 3. Effects of BBS on intracellular ROS levels in HDMECs stimulated with 200 μM H2O2. Cells were pre-treated with 200 μM AA, 2.5% BBS, or 2.5% BNB for 22 h, followed by exposure to 200 μM H2O2 for 2 h. (A) Representative DCF-DA fluorescence images showing intracellular ROS accumulation (green fluorescence, FITC channel; Fluorescein Isothiocyanate) and DIC (Differential Interference Contrast). Scale bar = 50 μm. Images are representative of three independent experiments. (B) Quantitative analysis of relative fluorescence intensity was performed using ImageJ software (version 1.53e). ‘+’ and ‘-’ indicate the presence and absence of the indicated substance, respectively. Data are presented as mean ± SD (n = 4). *** p < 0.005 relative to the H2O2-treated group. #### p < 0.0001 compared with the control group. No significant difference was observed between the BBS- and BNB-treated groups.
Figure 3. Effects of BBS on intracellular ROS levels in HDMECs stimulated with 200 μM H2O2. Cells were pre-treated with 200 μM AA, 2.5% BBS, or 2.5% BNB for 22 h, followed by exposure to 200 μM H2O2 for 2 h. (A) Representative DCF-DA fluorescence images showing intracellular ROS accumulation (green fluorescence, FITC channel; Fluorescein Isothiocyanate) and DIC (Differential Interference Contrast). Scale bar = 50 μm. Images are representative of three independent experiments. (B) Quantitative analysis of relative fluorescence intensity was performed using ImageJ software (version 1.53e). ‘+’ and ‘-’ indicate the presence and absence of the indicated substance, respectively. Data are presented as mean ± SD (n = 4). *** p < 0.005 relative to the H2O2-treated group. #### p < 0.0001 compared with the control group. No significant difference was observed between the BBS- and BNB-treated groups.
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Figure 4. Effect of BBS on mitochondrial membrane potential in HDMECs stimulated with 200 μM H2O2. Cells were pre-treated with 200 μM AA, 2.5% BBS, or 2.5% BNB for 22 h, followed by exposure to 200 μM H2O2 for 2 h. (A) Representative JC-1 fluorescence images showing green fluorescence representing depolarized mitochondria and red fluorescence indicating hyperpolarized mitochondria. MERGE indicates the merged images of RITC (Rhodamine isothiocyanate) and FITC (scale bar, 50 µm). Images represent results from three independent experiments. (B) Mitochondrial membrane potential was quantified using ImageJ software, version 1.53e. ‘+’ and ‘-’ indicate the presence and absence of the indicated substance, respectively. Data are presented as mean ± SD (n = 3), with statistical significance denoted as * p < 0.05, ** p < 0.01 relative to the H2O2-treated group. #### p < 0.0001 compared with the control group. No significant difference was observed between the BBS- and BNB-treated groups.
Figure 4. Effect of BBS on mitochondrial membrane potential in HDMECs stimulated with 200 μM H2O2. Cells were pre-treated with 200 μM AA, 2.5% BBS, or 2.5% BNB for 22 h, followed by exposure to 200 μM H2O2 for 2 h. (A) Representative JC-1 fluorescence images showing green fluorescence representing depolarized mitochondria and red fluorescence indicating hyperpolarized mitochondria. MERGE indicates the merged images of RITC (Rhodamine isothiocyanate) and FITC (scale bar, 50 µm). Images represent results from three independent experiments. (B) Mitochondrial membrane potential was quantified using ImageJ software, version 1.53e. ‘+’ and ‘-’ indicate the presence and absence of the indicated substance, respectively. Data are presented as mean ± SD (n = 3), with statistical significance denoted as * p < 0.05, ** p < 0.01 relative to the H2O2-treated group. #### p < 0.0001 compared with the control group. No significant difference was observed between the BBS- and BNB-treated groups.
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Figure 5. Effects of BBS on the phosphorylation levels of ERK, p38, and JNK in HDMECs stimulated with 200 μM H2O2. Cells were treated with 200 μM H2O2 followed by 200 μM AA, 2.5% BBS, or 2.5% BNB for 30 min. (A) Representative Western blot images showing the relative expression levels of each protein. (B) Relative expression levels of each protein. ‘+’ and ‘-‘ indicate the presence and absence of the indicated substance, respectively. Data are presented as mean ± SD (n = 3), with statistical significance denoted as * p < 0.05, ** p < 0.01 relative to the H2O2-treated group. # p < 0.05, and ## p < 0.01 compared with the control group. †† p < 0.01 compared with the BBS-treated group.
Figure 5. Effects of BBS on the phosphorylation levels of ERK, p38, and JNK in HDMECs stimulated with 200 μM H2O2. Cells were treated with 200 μM H2O2 followed by 200 μM AA, 2.5% BBS, or 2.5% BNB for 30 min. (A) Representative Western blot images showing the relative expression levels of each protein. (B) Relative expression levels of each protein. ‘+’ and ‘-‘ indicate the presence and absence of the indicated substance, respectively. Data are presented as mean ± SD (n = 3), with statistical significance denoted as * p < 0.05, ** p < 0.01 relative to the H2O2-treated group. # p < 0.05, and ## p < 0.01 compared with the control group. †† p < 0.01 compared with the BBS-treated group.
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Figure 6. Effects of BBS on tube formation in HDMECs stimulated with 200 μM H2O2. Cells were treated with 200 μM H2O2 followed by 200 μM AA, 2.5% BBS, or 2.5% BNB for 24 h. (A) Representative phase-contrast images showing tube formation in HDMECs at 6 and 24 h (scale bar, 50 µm). Images represent results from three independent experiments. (B,C) Quantitative analysis of tube formation parameters, including total tube length and number of nodes, was performed using ImageJ software, version 1.53e. ‘+’ and ‘-’ indicate the presence and absence of the indicated substance, respectively. (D) VEGFA secretion levels in HDMECs were quantified by ELISA after 6 h of treatment. Data are presented as mean ± SD (n = 3), with statistical significance denoted as * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 relative to the H2O2-treated group. # p < 0.05, #### p < 0.0001 compared with the control group. †† p < 0.01, ††† p < 0.001 compared with the BBS-treated group.
Figure 6. Effects of BBS on tube formation in HDMECs stimulated with 200 μM H2O2. Cells were treated with 200 μM H2O2 followed by 200 μM AA, 2.5% BBS, or 2.5% BNB for 24 h. (A) Representative phase-contrast images showing tube formation in HDMECs at 6 and 24 h (scale bar, 50 µm). Images represent results from three independent experiments. (B,C) Quantitative analysis of tube formation parameters, including total tube length and number of nodes, was performed using ImageJ software, version 1.53e. ‘+’ and ‘-’ indicate the presence and absence of the indicated substance, respectively. (D) VEGFA secretion levels in HDMECs were quantified by ELISA after 6 h of treatment. Data are presented as mean ± SD (n = 3), with statistical significance denoted as * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 relative to the H2O2-treated group. # p < 0.05, #### p < 0.0001 compared with the control group. †† p < 0.01, ††† p < 0.001 compared with the BBS-treated group.
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Figure 7. Schematic illustration of the protective role of BBS in HDMECs. H2O2 induced intracellular ROS accumulation, leading to cellular damage. BBS reduces ROS levels, restores damaged mitochondrial function, inhibits MAPK activation, and enhances angiogenic responses such as migration and tube formation in HDMECs, thereby preserving endothelial functionality and microvascular integrity under oxidative stress. (created with BioRender, https://www.biorender.com/, accessed on 8 January 2025).
Figure 7. Schematic illustration of the protective role of BBS in HDMECs. H2O2 induced intracellular ROS accumulation, leading to cellular damage. BBS reduces ROS levels, restores damaged mitochondrial function, inhibits MAPK activation, and enhances angiogenic responses such as migration and tube formation in HDMECs, thereby preserving endothelial functionality and microvascular integrity under oxidative stress. (created with BioRender, https://www.biorender.com/, accessed on 8 January 2025).
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MDPI and ACS Style

Shin, J.U.; Jo, Y.H.; Jeong, S.A.; Jeong, Y.H.; Son, M.G.; Jeong, Y.J.; Lim, B.O.; Shin, D.W. Bioconverted Blueberry Extract Potentiates the Angiogenic and Endothelial Functions in Human Dermal Microvascular Endothelial Cells Under Oxidative Stress. Curr. Issues Mol. Biol. 2026, 48, 224. https://doi.org/10.3390/cimb48020224

AMA Style

Shin JU, Jo YH, Jeong SA, Jeong YH, Son MG, Jeong YJ, Lim BO, Shin DW. Bioconverted Blueberry Extract Potentiates the Angiogenic and Endothelial Functions in Human Dermal Microvascular Endothelial Cells Under Oxidative Stress. Current Issues in Molecular Biology. 2026; 48(2):224. https://doi.org/10.3390/cimb48020224

Chicago/Turabian Style

Shin, Jung Un, Yun Hoo Jo, Soo Ah Jeong, Yeong Hwan Jeong, Myeong Gwan Son, Yoo Jeong Jeong, Beong Ou Lim, and Dong Wook Shin. 2026. "Bioconverted Blueberry Extract Potentiates the Angiogenic and Endothelial Functions in Human Dermal Microvascular Endothelial Cells Under Oxidative Stress" Current Issues in Molecular Biology 48, no. 2: 224. https://doi.org/10.3390/cimb48020224

APA Style

Shin, J. U., Jo, Y. H., Jeong, S. A., Jeong, Y. H., Son, M. G., Jeong, Y. J., Lim, B. O., & Shin, D. W. (2026). Bioconverted Blueberry Extract Potentiates the Angiogenic and Endothelial Functions in Human Dermal Microvascular Endothelial Cells Under Oxidative Stress. Current Issues in Molecular Biology, 48(2), 224. https://doi.org/10.3390/cimb48020224

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