Boron-Enhanced Mitochondrial Repair: DeepA-I Tissue Regeneration

Abstract

Cellular metabolism is a key regulator of tissue repair and regeneration, with mitochondrial function playing a central role in energy production and cellular homeostasis. Dysfunctional mitochondria, often due to excessive reactive oxygen species (ROS), contribute to oxidative stress, impaired wound healing, and chronic inflammation. This study investigates the therapeutic potential of DeepA-I, a Boron-enriched compound, in enhancing mitochondrial health, reducing oxidative damage, and promoting cellular repair in human umbilical vein endothelial cells (HUVEC) and mouse embryonic fibroblasts (MEF). Boron quantification via inductively coupled plasma optical emission spectroscopy (ICP-OES) confirmed its presence in DeepA-I. Cytotoxicity assessment (MTT assay) demonstrated its safety, while fluorescence microscopy (DAPI, MitoSPY, DCFDA) revealed reduced ROS levels and preserved mitochondrial integrity. A scratch assay showed accelerated cell migration and wound closure in DeepA-I-treated cells. Western blot analysis indicated the downregulation of Akt (a proliferation marker) and the upregulation of NRF2, a key regulator of oxidative stress resistance, without affecting apoptosis-related proteins. These results suggest that DeepA-I, via its Boron-mediated mechanisms, enhances mitochondrial function, mitigates ROS-induced damage, and improves tissue repair, positioning it as a promising therapeutic candidate for inflammatory and degenerative conditions.

Keywords: Boron supplementation; oxidative stress; reactive oxygen species (ROS); tissue repair.

Figure 1

Dose-dependent MTT results for DeepA-I in MEF (a) and HUVEC (b) cells. Error bars indicate standard deviations of 3 biological and 4 technical replicates. For control groups, cells were treated with a medium without DeepA-I. ns. p > 0.05.

Figure 2

(a) Representation of mitochondria (MitoSPY, red) and cell nuclei (DAPI, blue) after 24 h of DeepA-I treatment (1:10, 1:100, 1:1000, 1:5000, and 1:10,000) in MEF cells. The representative image is selected from 2 repetitive images. The scale bar is 150 μm. (b) Quantitative analysis of mitochondrial fluorescence under control and DeepA-I-treated conditions. Data are represented as mean ± SEM (^∗∗∗p < 0.001, ^∗∗p < 0.01, ^∗p < 0.05).

Figure 3

(a) Representative images of mitochondria (MitoSPY, red), cell nuclei (DAPI, blue), and ROS activity (DCFDA) following DeepA-I treatment 24 h (1:10, 1:100, 1:1000, 1:5000, and 1:10,000) in HUVEC cells. The representative image is selected from 2 repetitive images. The scale bar is 150 μm. (b) Quantitative analysis of mitochondrial fluorescence under control and DeepA-I-treated conditions. (c) Quantitative analysis of ROS level under control and DeepA-I-treated conditions. Data are represented as mean ± SEM (^∗∗∗∗p < 0.0001, ^∗∗p < 0.01, ^∗p < 0.05).

Figure 4

Wound healing-migration analysis was performed in HUVEC and MEF cells following two DeepA-I treatments. Microscopic images of HUVEC cell migration were taken at 0, 24, and 48 h. The same analysis was done at only 24 h exposure of MEF cells. Wound closure (%) in both cell lines was analysed to track the migration profile of the cells in the presence or absence of DeepA-I. Representative images were selected from 2 biological replicates.

Figure 5

Analysis of the changes in protein expression by immunoblotting. Cell survival, antioxidant capacity, apoptosis, and autophagy pathways were analyzed at the protein level in HUVEC (a) and MEF (b) cells with and without DeepA-I treatment. β-actin was used as a control. Representative images were selected from 2 biological replicates.