Fig.3 Model of laser treatment in wound of rats. (A) laser automated
surgical platform with different incident angles. B) Schematic
cross-sectional view of the treatment process of rat skin tissue. C)
Schematic of laser scanning
path-Zigzag .
We
monitored and quantitatively measured the wound healing process at
pre-determined intervals post-operation as depicted in Fig.4(A). The
observations revealed innate self-healing capabilities in rats as
evidenced by the continuous reduction in wound area in the control
group, despite wounds remaining unhealed after 24 days without any
additional treatment. It appears that low-energy laser treatment at
1064nm can foster wound healing to an extent, as all three laser-treated
groups exhibited smaller wound areas compared to the control group at
every time-point post-treatment. Notably, complete recovery was achieved
in all laser-treated groups within 14 days, marking a healing period
41.7% shorter than that of the control samples. As per Fig.4(C) and
Fig.4(B), the 30°Laser samples showed the quickest healing speed during
the first stage (initial 3 days) of laser welding, while the 90°Laser
samples exhibited the fastest healing speed in the second stage. For the
60°Laser samples, even though the initial healing speed wasn’t
particularly prominent, the healing processes post the second laser weld
were the most significant. It remains challenging to determine the
optimal parameter group based solely on the trend of healing speed and
wound size since factors like collagen fiber disposition, distribution
of inflammatory factors, and varying types of collagen content are all
vital indicators of healing performance. To further scrutinize the
differences between incident angles and the mechanism of laser welding
tissue, we proceeded with comprehensive characterization and analysis of
cells, collagen, and functional protein groups during the healing
process. This data has been incorporated into Fig.4(C) and will be
detailed in the following sections.
Fig.4 Whole process of wound healing. (A)
Representative images of macro
appearances of rats during whole healing processes in three groups. (B)
Schematic diagram of the wounds managed by different laser treatments in
24 days. (C) Quantitative data of wound sizes at different time points
(n ¼ 3). (D) Quantitative data of wound healing ratio at different time
points (n ¼ 3). Data represent mean \(\pm\) SD; *, P < 0.05, **, P <
0.01.
Microstructures of rats
after
laser welding with different incident angles
In trauma treatment, the wound healing process is typically segmented
into stages: the inflammatory response stage, the granulation stage, and
the epithelial formation stage. These stages often overlap with each
other. Figure 5 presents Hematoxylin and Eosin (H&E) stained images of
the wound tissue at specific time points, which include days 3, 7, and
14. The figure distinctly reveals noticeable differences in the
microstructure of samples throughout the healing process under varied
laser incident angles.
On day 3, post the first laser welding, the incision location and gap
are still clearly visible at 200 μm magnification. Following the first
laser weld, the most pronounced recovery was observed in the 90° laser
and 30° laser groups; the wound incision gap in the dermis layer was
considerably reduced, and partial remodeling of the collagen in the
dermis had occurred through integrin interactions. On the contrary, the
samples in the 60° group exhibited significant epidermal gaps and it
remained challenging to discern clear collagen deposition in the dermis.
Following the second laser welding on day 3, marked improvements were
noted across all groups, with particularly tremendous healing progress
in both the 90° and 30° laser groups. At this stage, the incision’s
micromorphology was barely noticeable in all three groups, and
re-epithelialization of all samples had commenced along with new
granulation formation. This re-epithelialization process is pivotal to
accelerating dermal healing; the beneficial thickening of the upper
epidermis serves as a bio-functional barrier established early in the
wound healing process. This barrier plays a crucial role in preventing
excessive transdermal water loss and further wound infection.
By the 14th day of the healing process, it was observed that any
existing gaps in the wound incision had entirely disappeared. The upper
epidermis was completely covered, and a dense new collagen network
structure was evident in the dermis.