中华烧伤与创面修复杂志

National expert consensus on prone position therapy in adult burn patients (2022 version)

The Burn and Trauma Branch of Chinese Geriatrics Society, Critical Care Group of Chinese Burn Association

2022, 38(7): 601-609. doi: 10.3760/cma.j.cn501120-20211208-00407

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Prone position ventilation (PPV) is attracting more and more attention as a part of mechanical ventilation treatment, and relevant consensus and guidelines have been formulated. Prone position therapy (PPT) is widely used in the clinical diagnosis and treatment of burns. Compared with traditional PPV, burn PPT is significantly different in indications, process details, precautions, etc. Therefore, the Burn and Trauma Branch of Chinese Geriatric Society and Critical Care Group of Chinese Burn Association collected the evidence,led the formulation of National expert consensus on prone position therapy in adult burn patients (2022 version), and formulated recommendations on action mechanism, indications, use process of PPT for clinical guidance.

Progress and thoughts on the regulation of wound repair by growth factors

Xiao Jian, Zhang Fan

2022, 38(7): 610-615. doi: 10.3760/cma.j.cn501225-20220416-00139

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Growth factors play an important role in wound healing, and they mainly accelerate wound healing by activating the related signal pathways. Chinese scientists have been conducting basic and clinical researches on growth factors for 30 years, with a series of growth factor drugs being developed and widely used in the treatment of burns and trauma and chronic refractory ulcers. This paper expounds the frontier progress of growth factors on wound healing from the perspectives of immunity, nerve, fat, and so on, and puts forward the further thoughts of the research team on the regulation of wound healing by growth factors.

Effects of in situ cross-linked graphene oxide-containing gelatin methacrylate anhydride hydrogel on wound vascularization of full-thickness skin defect in mice

Liang Liting, Song Wei, Zhang Chao, Li Zhao, Yao Bin, Zhang Mengde, Yuan Xingyu, Enhejirigala, Fu Xiaobing, Huang Sha, Zhu Ping

2022, 38(7): 616-628. doi: 10.3760/cma.j.cn501225-20220314-00063

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Objective To prepare graphene oxide (GO)-containing gelatin methacrylate anhydride (GelMA) hydrogel and to investigate the effects of in situ photopolymerized GO-GelMA composite hydrogel in wound vascularization of full-thickness skin defect in mice. Methods The experimental study method was used. The 50 μL of 0.2 mg/mL GO solution was evenly applied onto the conductive gel, and the structure and size of GO were observed under field emission scanning electron microscope after drying. Human skin fibroblasts (HSFs) were divided into 0 μg/mL GO (without GO solution, the same as below) group, 0.1 μg/mL GO group, 1.0 μg/mL GO group, 5.0 μg/mL GO group, and 10.0 μg/mL GO group treated with GO of the corresponding final mass concentration, and the absorbance value was detected using a microplate analyzer after 48 h of culture to reflect the proliferation activity of cells (n=6). HSFs and human umbilical vein vascular endothelial cells (HUVECs) were divided into 0 μg/mL GO group, 0.1 μg/mL GO group, 1.0 μg/mL GO group, and 5.0 μg/mL GO group treated with GO of the corresponding final mass concentration, and the migration rates of HSFs at 24 and 36 h after scratching (n=5) and HUVECs at 12 h after scratching (n=3) were detected by scratch test, and the level of vascular endothelial growth factor (VEGF) secreted by HSFs after 4, 6, and 8 h of culture was detected by enzyme-linked immunosorbent assay method (n=3). The prepared GO-GelMA composite hydrogels containing GO of the corresponding final mass concentration were set as 0 μg/mL GO composite hydrogel group, 0.1 μg/mL GO composite hydrogel group, 1.0 μg/mL GO composite hydrogel group, and 5.0 μg/mL GO composite hydrogel group to observe their properties before and after cross-linking, and to detect the release of GO after soaking with phosphate buffer solution for 3 and 7 d (n=3). The full-thickness skin defect wounds were made on the back of 16 6-week-old female C57BL/6 mice. The mice treated with in situ cross-linked GO-GelMA composite hydrogel containing GO of the corresponding final mass concentration were divided into 0 μg/mL GO composite hydrogel group, 0.1 μg/mL GO composite hydrogel group, 1.0 μg/mL GO composite hydrogel group, and 5.0 μg/mL GO composite hydrogel group according to the random number table, with 4 mice in each group. The general condition of wound was observed and the wound healing rate was calculated on 3, 7, and 14 d of treatment, the wound blood perfusion was detected by laser Doppler flowmetry on 3, 7, and 14 d of treatment and the mean perfusion unit (MPU) ratio was calculated, and the wound vascularization on 7 d of treatment was observed after hematoxylin-eosin staining and the vascular density was calculated (n=3). The wound tissue of mice in 0 μg/mL GO composite hydrogel group and 0.1 μg/mL GO composite hydrogel group on 7 d of treatment was collected to observe the relationship between the distribution of GO and neovascularization by hematoxylin-eosin staining (n=3) and the expression of VEGF by immunohistochemical staining. Data were statistically analyzed with analysis of variance for repeated measurement, one-way analysis of variance, and Tukey's method. Results GO had a multilayered lamellar structure with the width of about 20 μm and the length of about 50 μm. The absorbance value of HSFs in 10.0 μg/mL GO group was significantly lower than that in 0 μg/mL GO group after 48 h of culture (q=7.64, P<0.01). At 24 h after scratching, the migration rates of HSFs were similar in the four groups (P>0.05); at 36 h after scratching, the migration rate of HSFs in 0.1 μg/mL GO group was significantly higher than that in 0 μg/mL GO group, 1.0 μg/mL GO group, and 5.0 μg/mL GO group (with q values of 7.48, 10.81, and 10.20, respectively, P<0.01). At 12 h after scratching, the migration rate of HUVECs in 0.1 μg/mL GO group was significantly higher than that in 0 μg/mL GO group, 1.0 μg/mL GO group, and 5.0 μg/mL GO group (with q values of 7.11, 8.99, and 14.92, respectively, P<0.01), and the migration rate of HUVECs in 5.0 μg/mL GO group was significantly lower than that in 0 μg/mL GO group and 1.0 μg/mL GO group (with q values of 7.81 and 5.33, respectively, P<0.05 or P<0.01 ). At 4 and 6 h of culture, the VEGF expressions of HSFs in the four groups were similar (P>0.05); at 8 h of culture, the VEGF expression of HSFs in 0.1 μg/mL GO group was significantly higher than that in 0 μg/mL GO group and 5.0 μg/mL GO group (with q values of 4.75 and 4.48, respectively, P<0.05). The GO-GelMA composite hydrogels in the four groups were all red liquid before cross-linking, which turned to light yellow gel after cross-linking, with no significant difference in fluidity. The GO in the GO-GelMA composite hydrogel of 0 μg/mL GO composite hydrogel group had no release of GO at all time points; the GO in the GO-GelMA composite hydrogels of the other 3 groups was partially released on 3 d of soaking, and all the GO was released on 7 d of soaking. From 3 to 14 d of treatment, the wounds of mice in the 4 groups were covered with hydrogel dressings, kept moist, and gradually healed. On 3, 7, and 14 d of treatment, the wound healing rates of mice in the four groups were similar (P>0.05). On 3 d of treatment, the MPU ratio of wound of mice in 0.1 μg/mL GO composite hydrogel group was significantly higher than that in 0 μg/mL GO composite hydrogel group, 1.0 μg/mL GO composite hydrogel group, and 5.0 μg/mL GO composite hydrogel group (with q values of 10.70, 11.83, and 10.65, respectively, P<0.05 or P<0.01). On 7 and 14 d of treatment, the MPU ratios of wound of mice in the four groups were similar (P>0.05). The MPU ratio of wound of mice in 0.1 μg/mL GO composite hydrogel group on 7 d of treatment was significantly lower than that on 3 d of treatment (q=14.38, P<0.05), and that on 14 d of treatment was significantly lower than that on 7 d of treatment (q=27.78, P<0.01). On 7 d of treatment, the neovascular density of wound of mice on 7 d of treatment was 120.7±4.1 per 200 times of visual field, which was significantly higher than 61.7±1.3, 77.7±10.2, and 99.0±7.9 per 200 times of visual field in 0 μg/mL GO composite hydrogel group, 1.0 μg/mL GO composite hydrogel group, and 5.0 μg/mL GO composite hydrogel group (with q values of 12.88, 7.79, and 6.70, respectively, P<0.01), and the neovascular density of wound of mice in 1.0 μg/mL GO composite hydrogel group and 5.0 μg/mL GO composite hydrogel group was significantly higher than that in 0 μg/mL GO composite hydrogel group (with q values of 5.10 and 6.19, respectively, P<0.05). On 7 d of treatment, cluster of new blood vessels in wound of mice in 0.1 μg/mL GO composite hydrogel group was significantly more than that in 0 μg/mL GO composite hydrogel group, and the new blood vessels were clustered near the GO; a large amount of VEGF was expressed in wound of mice in 0.1 μg/mL GO composite hydrogel group in the distribution area of GO and new blood vessels. Conclusions GO with mass concentration lower than 10.0 μg/mL had no adverse effect on proliferation activity of HSFs, and GO of 0.1 μg/mL can promote the migration of HSFs and HUVECs, and can promote the secretion of VEGF in HSFs. In situ photopolymerized of GO-GelMA composite hydrogel dressing can promote the wound neovascularization of full-thickness skin defect in mice and increase wound blood perfusion in the early stage, with GO showing an enrichment effect on angiogenesis, and the mechanism may be related to the role of GO in promoting the secretion of VEGF by wound cells.

Investigation on the growth factor regulatory network of dermal fibroblasts in mouse full-thickness skin defect wounds based on single-cell RNA sequencing

Sun Lixiang, Wu Shuai, Zhang Xiaowei, Liu Wenjie, Zhang Lingjuan

2022, 38(7): 629-639. doi: 10.3760/cma.j.cn501225-20220215-00029

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Objective To explore the heterogeneity and growth factor regulatory network of dermal fibroblasts (dFbs) in mouse full-thickness skin defect wounds based on single-cell RNA sequencing. Methods The experimental research methods were adopted. The normal skin tissue from 5 healthy 8-week-old male C57BL/6 mice (the same mouse age, sex, and strain below) was harvested, and the wound tissue of another 5 mice with full-thickness skin defect on the back was harvested on post injury day (PID) 7. The cell suspension was obtained by digesting the tissue with collagenase D and DNase Ⅰ, sequencing library was constructed using 10x Genomics platform, and single-cell RNA sequencing was performed by Illumina Novaseq6000 sequencer. The gene expression matrices of cells in the two kinds of tissue were obtained by analysis of Seurat 3.0 program of software R4.1.1, and two-dimensional tSNE plots classified by cell group, cell source, and gene labeling of major cells in skin were used for visual display. According to the existing literature and the CellMarker database searching, the expression of marker genes in the gene expression matrices of cells in the two kinds of tissue was analyzed, and each cell group was numbered and defined. The gene expression matrices and cell clustering information were introduced into CellChat 1.1.3 program of software R4.1.1 to analyze the intercellular communication in the two kinds of tissue and the intercellular communication involving vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), epidermal growth factor (EGF), and fibroblast growth factor (FGF) signal pathways in the wound tissue, the relative contribution of each pair of FGF subtypes and FGF receptor (FGFR) subtypes (hereinafter referred to as FGF ligand receptor pairs) to FGF signal network in the two kinds of tissue, and the intercellular communication in the signal pathway of FGF ligand receptor pairs with the top 2 relative contributions in the two kinds of tissue. The normal skin tissue from one healthy mouse was harvested, and the wound tissue of one mouse with full-thickness skin defect on the back was harvested on PID 7. The multiple immunofluorescence staining was performed to detect the expression and distribution of FGF7 protein and its co-localized expression with dipeptidyl peptidase 4 (DPP4), stem cell antigen 1 (SCA1), smooth muscle actin (SMA), and PDGF receptor α (PDGFRα) protein. Results Both the normal skin tissue of healthy mice and the wound tissue of full-thickness skin defected mice on PID 7 contained 25 cell groups, but the numbers of cells in each cell group between the two kinds of tissue were different. Genes PDGFRα, platelet endothelial cell adhesion molecule 1, lymphatic endothelial hyaluronic acid receptor 1, receptor protein tyrosine phosphatase C, keratin 10, and keratin 79 all had distinct distributions on two-dimensional tSNE plots, indicating specific cell groups respectively. The 25 cell groups were numbered by C0-C24 and divided into 9 dFb subgroups and 16 non-dFb groups. dFb subgroups included C0 as interstitial progenitor cells, C5 as adipose precursor cells, and C13 as contractile muscle cells related fibroblasts, etc. Non-dFb group included C3 as neutrophils, C8 as T cells, and C18 as erythrocytes, etc. Compared with that of the normal skin tissue of healthy mice, the intercellular communication in the wound tissue of full-thickness skin defected mice on PID 7 was more and denser, and the top 3 cell groups in intercellular communication intensity were dFb subgroups C0, C1, and C2, of which all communicated with other cell groups in the wound tissue. In the wound tissue of full-thickness skin defected mice on PID 7, VEGF signals were mainly sent by the dFb subgroup C0 and received by vascular related cell groups C19 and C21, PDGF signals were mainly sent by peripheral cells C14 and received by multiple dFb subgroups, EGF signals were mainly sent by keratinocyte subgroups C9 and C11 and received by the dFb subgroup C0, and the main sender and receiver of FGF signals were the dFb subgroup C6. In the relative contribution rank of FGF ligand receptor pairs to FGF signal network in the normal skin tissue of healthy mice and the wound tissue of full-thickness skin defected mice on PID 7, FGF7-FGFR1 was the top 1, and FGF7-FGFR2 or FGF10-FGFR1 was in the second place, respectively; compared with those in the normal skin tissue, there was more intercellular communication in FGF7-FGFR1 signal pathway, while the intercellular communication in FGF7-FGFR2 and FGF10-FGFR1 signal pathways decreased slightly or did not change significantly in the wound tissue; the intercellular communication in FGF7-FGFR1 signal pathway in the wound tissue was stronger than that in FGF7-FGFR2 or FGF10-FGFR1 signal pathway; in the two kinds of tissue, FGF7 signal was mainly sent by dFb subgroups C0, C1, and C2, and received by dFb subgroups C6 and C7. Compared with that in the normal skin tissue of healthy mouse, the expression of FGF7 protein was higher in the wound tissue of full-thickness skin defected mouse on PID 7; in the normal skin tissue, FGF7 protein was mainly expressed in the skin interstitium and also expressed in the white adipose tissue near the dermis layer; in the two kinds of tissue, FGF7 protein was co-localized with DPP4 and SCA1 proteins and expressed in the skin interstitium, co-localized with PDGFRα protein and expressed in dFbs, but was not co-localized with SMA protein, with more co-localized expression of FGF7 in the wound tissue than that in the normal skin tissue. Conclusions In the process of wound healing of mouse full-thickness skin defect wound, dFbs are highly heterogeneous, act as potential major secretory or receiving cell populations of a variety of growth factors, and have a close and complex relationship with the growth factor signal pathways. FGF7-FGFR1 signal pathway is the main FGF signal pathway in the process of wound healing, which targets and regulates multiple dFb subgroups.

Effects of methacrylic anhydride gelatin hydrogel loaded with silver and recombinant human basic fibroblast growth factor on deep partial-thickness burn wounds in rabbits

Chen Xiangjun, Wu Xing, Lin Huanhuan, Liu Zhaoxing, Liu Sha

2022, 38(7): 640-649. doi: 10.3760/cma.j.cn501120-20210726-00260

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Objective To investigate the effects of methacrylic anhydride gelatin (GelMA) hydrogel loaded with silver and recombinant human basic fibroblast growth factor (rh-bFGF) on deep partial-thickness burn wounds in rabbits. Methods The experimental research method was adopted. Low-concentration GelMA materials, medium-concentration GelMA materials and high-concentration GelMA materials containing different concentrations of methacrylic anhydride (MA) were prepared, after adding photoinitiator, low-concentration GelMA hydrogels, medium-concentration GelMA hydrogels, and high-concentration GelMA hydrogels were obtained, respectively. The nuclear magnetic resonance spectroscopy was performed to detect the hydrogen nuclear magnetic resonance spectra of the above-mentioned three concentrations of GelMA materials, and to calculate the degree of substitution according to the spectrum diagram. The three-dimensional microstructure and pore size of 3 types of above-mentioned GelMA hydrogels were detected by field emission scanning electron microscopy (FESEM), with 9 samples measured. According to the selected concentration of MA, ten kinds of solutions of GelMA with different concentration of silver (silver-containing GelMA) were synthesized, and the silver-containing GelMA solution of each concentration was divided into three parts, and then exposed to ultraviolet light lasting for 20, 25, and 35 s, respectively. After adding photoinitiator,the corresponding silver-containing GelMA hydrogels were obtained. The residual degradation rate of silver-containing GelMA hydrogel with different photocrosslinking times was detected by collagenase degradation method at degradation of 12, 24, 36, and 48 h; and the time required for complete degradation was detected, and the sample number was 5. The inhibition zone diameter of GelMA hydrogel under above screened photocrosslinking times containing 10 concentrations of silver against Staphylococcus aureus was measured to reflect its antibacterial ability, and the sample numbers were all 5. The silver-containing GelMA hydrogel with statistical significance compared with the antibacterial circle diameter of the silver-containing GelMA hydrogel containing the lowest concentration (no silver) was considered as having antibacterial activity. The three-dimensional microstructure and pore size of the silver-containing GelMA hydrogels with antibacterial activity and the lowest drug concentration selected were detected by FESEM, and the sample numbers were all 9. The freeze-dried alone GelMA hydrogel and the freeze-dried silver-containing GelMA hydrogel were soaked in phosphate buffer solution for 24 h, respectively, then the swelling rate of the two GelMA hydrogel were calculated and compared by weighing method, and the sample number was 5. GelMA hydrogel containing silver and rh-bFGF, namely compound hydrogel for short, was prepared according to the preliminary experiment and the above experimental results. The appearance of the composite hydrogel was observed in general, and its three-dimensional microstructure and pore size were detected by FESEM. The deep partial-thickness burn wound was made on the back of 30 rabbits (aged 4-6 months, female half and half). Meanwhile, with the rabbit head as the benchmark, the wounds on the left side of the spine were treated as composite hydrogel treatment group, and the wounds on the right side were treated as gauze control group, and which were treated accordingly. On post injury day (PID) 3, 7, 14, 21, and 28, the healing of wounds in the two groups was observed. On PID 7, 14, 21, and 28, the wound healing area was recorded and the healing rate was calculated, with a sample number of 30. Data were statistically analyzed with analysis of variance for repeated measurement, one-way analysis of variance, and independent sample t test. Results The substitution degree among low-concentration GelMA materials, medium-concentration GelMA materials, and high-concentration GelMA materials was significantly different (F=1 628.00, P<0.01). The low-concentration GelMA hydrogel had a loose and irregular three-dimensional spatial network structure with a pore size of (60±17) μm; the medium-concentration GelMA hydrogel had a relatively uniform three-dimensional spatial network and pore size with a pore size of (45±13) μm; the high-concentration GelMA hydrogel had the dense and disordered three-dimensional spatial network with a pore size of (25±15) μm, the pore sizes of 3 types of GelMA hydrogels were significantly differences (F=12.20, P<0.01), and medium concentration of MA was selected for the concentration of subsequent materials. The degradability of silver-containing GelMA hydrogels with different concentrations of the same photocrosslinking time was basically same. The degradation residual rates of silver-containing GelMA hydrogels with 20, 25, and 35 s crosslinking time at 12 h were (74.2±1.7)%, (85.3±0.9)%, and (93.2±1.2)%, respectively; the residual rates of degradation at 24 h were (58.3±2.1)%, (65.2±1.8)%, and (81.4±2.6)%, respectively; the residual rates of degradation at 36 h were (22.4±1.9)%, (45.2±1.7)%, and (68.1±1.4)%, respectively; the residual rates of degradation at 48 h were (8.2±1.7)%, (32.4±1.3)%, and (54.3±2.2)%, respectively, and 20, 25, and 30 s photocrosslinking time required for complete degradation of silver-containing GelMA hydrogels were (50.2±2.4), (62.4±1.4), and (72.2±3.2) h, and the difference was statistically significant (F=182.40, P<0.01), 25 s were selected as the subsequent photocrosslinking time. The antibacterial diameters of 10 types of silver-containing GelMA hydrogels against Staphylococcus aureus from low to high concentrations were (2.6±0.4), (2.5±0.4), (3.2±0.4), (12.1±0.7), (14.8±0.7), (15.1±0.5), (16.2±0.6), (16.7±0.5), (16.7±0.4), and (16.7±0.6) mm, respectively, and which basically showed a concentration-dependent increasing trend, and the overall difference was statistically significant (F=428.70, P<0.01). Compared with the silver-containing GelMA hydrogel with the lowest concentration, the antibacterial circle diameters of other silver-containing GelMA hydrogels with antibacterial ability from low to high concentration were significantly increased (with t values of 26.35, 33.84, 43.65, 42.17, 49.24, 55.74, and 43.72, respectively, P<0.01). The silver-containing GelMA hydrogel with the antibacterial diameter of (12.1±0.7) mm had the lowest antibacterial activity against Staphylococcus aureus and the lowest drug loading concentration, and the concentration of silver was selected for the concentration of subsequent materials. The microscopic morphology of the silver-containing GelMA hydrogel containing silver element with a pore size of (45±13) μm had a regular and linear strip-like structure. After soaking for 24 h, the swelling ratio of silver-containing GelMA hydrogel was similar to that of alone GelMA hydrogel. The composite hydrogel was colorless, clear and transparent, and its three-dimensional microstructure was a regular and uniform grid, with a filament network structure inside, and the pore size of (40±21) μm. On PID 3, a large amount of necrotic tissue and exudate of rabbit wound in composite hydrogel group were observed, and scattered scabs, a small amount of necrotic tissue and exudate of rabbit wound in gauze control group were observed. On PID 7, the area of rabbit wound in composite hydrogel group was significantly reduced, and adhesion of rabbit wound and gauze in gauze control group was observed. On PID 14, In composite hydrogel group, the rabbit wound surface was ruddy, and the growth of granulation tissue was observed, and in gauze control group, the rabbit wound base was pale, and the blood supply was poor. On PID 21, the rabbit wounds in composite hydrogel group healed completely, and rabbit wound in gauze control group had healing trend. On PID 28, new hair could be seen on rabbit wound surface in composite hydrogel group; oval wound of rabbit in gauze control group still remained. On PID 7, 14, 21, and 28, the wound healing areas of rabbit in composite hydrogel group were significantly larger than those in gauze control group (with t values of 2.24, 4.43, 7.67, and 7.69, respectively, P<0.05 or P<0.01). Conclusions The medium-concentration GelMA hydrogel has good physical and chemical properties in terms of swelling and degradability. The screened silver-containing GelMA hydrogels had the lowest antibacterial activity and the lowest drug loading concentration. Composite hydrogel can significantly shorten the healing time of deep partial-thickness burn wounds in rabbits.

Feasibility study on the preparation of novel negative pressure materials for constructing new matrix of full-thickness skin defect wounds in rats

Liu Yifan, Jiang Zhaoqi, Huang Yao, Ni Pengwen, Xie Ting

2022, 38(7): 650-660. doi: 10.3760/cma.j.cn501120-20210401-00113

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Objective To explore the feasibility on the preparation of novel negative pressure materials for constructing new matrix of full-thickness skin defect wounds in rats. Methods The experimental research method was applied. The microstructure of polyurethane foam dressing which was commonly used in negative pressure treatment was observed under scanning electron microscope, and its pore diameter was detected (n=5). Polycaprolactone (PCL) and polybutylene succinate (PBS) were used respectively as raw materials for the preparation of PCL and PBS negative pressure materials by melt spinning technology, with the measured pore diameter of polyurethane foam dressing as the spinning spacing at the spinning rates of 15, 25, and 35 mm/s, respectively. The microstructures of the prepared negative pressure materials were observed under scanning electron microscope, and their fiber diameters were measured. The tensile strength and tensile modulus of the prepared negative pressure materials and polyurethane foam dressing were measured by tensile testing machine and composite testing machine, respectively (n=5), to screen the spinning rate for subsequent preparation of negative pressure materials. Human skin fibroblasts (Fbs) in logarithmic growth phase were co-cultured with PCL negative pressure material and PBS negative pressure material prepared at the selected spinning rate, respectively. After 1, 4, and 7 day (s) of co-culture, the cell activity and adhesion in the materials was detected by living/dead cells detection kit, and the cell proliferation level in the materials was detected by cell counting kit 8 method (n=5). A full-thickness skin defect wound was prepared on the back of 18 5-6 weeks old Sprague-Dawley rats (gender unlimited). Immediately after injury, the injured rats were divided into PCL+polyurethane group, PBS+polyurethane group, and polyurethane alone group according to the random number table (with 6 rats in each group). The wounds were covered with materials containing corresponding component and performed with continuous negative pressure suction at the negative pressure of -16.7 kPa. The wound tissue along with materials directly contacted to the wound (hereinafter referred to as wound specimens) were collected from 3 rats in each group after 7 and 14 days of negative pressure treatment (NPT), respectively. The growth of granulation tissue and the attachment of material to wound surface were observed after hematoxylin-eosin staining, the collagen fiber deposition was observed after Masson staining, and CD34 and interleukin-6 (IL-6) positive cells were detected and counted by immunohistochemical staining. Data were statistically analyzed with one-way analysis of variance, analysis of variance for factorial design, least significant difference-t test, Kruskal-Wallis H test, Mann-Whitney U test, and Bonferroni correction. Results The microstructure of polyurethane foam dressing was loose and porous, with the pore diameter of (815±182) μm. The spinning spacing for the subsequent negative pressure material was set as 800 μm. The microstructures of PBS negative pressure material and PCL negative pressure material were regular, with vertically interconnected layers and continuous fibers in even thickness, but the fibers of PBS negative pressure material were straighter than those of PCL negative pressure material. There was no obvious difference in the microstructure of negative pressure materials prepared from the same raw material at different spinning rates. The fiber diameters of PCL negative pressure materials prepared at three spinning rates were similar (P>0.05). The fiber diameters of PBS negative pressure materials prepared at spinning rates of 25 mm/s and 35 mm/s were significantly smaller than the fiber diameter of PBS negative pressure material prepared at the spinning rate of 15 mm/s (with t values of 4.99 and 6.40, respectively, P<0.01). Both the tensile strength and tensile modulus of PCL negative pressure materials prepared at three spinning rates were similar (P>0.05). The tensile strength of PBS negative pressure materials prepared at spinning rates of 15 mm/s and 25 mm/s was significantly lower than that of PBS negative pressure materials prepared at the spinning rate of 35 mm/s (with t values of 9.20 and 8.92, respectively, P<0.01), and the tensile modulus was significantly lower than that of PBS negative pressure materials prepared at the spinning rate of 35 mm/s (with t values of 2.58 and 2.47, respectively, P<0.05). Subsequently, PCL negative pressure material was prepared at the spinning rate of 35 mm/s, and PBS negative pressure material was prepared at the spinning rate of 15 mm/s. After 1, 4, and 7 day (s) of co-culture, the number of human skin Fbs that adhered to PCL negative pressure material and PBS negative pressure material increased with time, and there was no significant difference between the two materials. After 1 and 7 day (s) of co-culture, the proliferation levels of human skin Fbs between the two negative pressure materials were similar (P>0.05). After being co-cultured for 4 days, the proliferation level of human skin Fbs in PBS negative pressure material was significantly higher than that in PCL negative pressure material (t=6.37, P<0.01). After 7 days of NPT, the materials were clearly identifiable and a small amount of collagen fibers were also observed in the wound specimens of rats in the three groups; a small amount of granulation tissue was observed in the wound specimens of rats in polyurethane alone group. After 14 days of NPT, a large number of granulation tissue and collagen fibers were observed in the wound specimens of rats in the three groups; the materials and wound tissue in the wound specimens of rats in PCL+polyurethane group could not be clearly distinguished. After 7 and 14 days of NPT, the collagen fibers in the wound specimens of rats in polyurethane alone group were denser than those in the other two groups. After 7 days of NPT, the number of CD34 positive cells in the wound specimens of rats in PBS+polyurethane group was 14.8±3.6 per 400 times visual field, which was significantly less than 27.8±9.1 in polyurethane alone group (t=3.06, P<0.05); the number of IL-6 positive cells was 60 (49, 72), which was significantly more than 44 (38, 50) in polyurethane alone group (Z=2.41, P<0.05). After 14 days of NPT, the number of IL-6 positive cells in the wound specimens of rats in PBS+polyurethane group was 19 (12, 28) per 400 times visual field, which was significantly more than 3 (1, 10) in PCL+polyurethane group and 9 (2, 13) in polyurethane alone group (with Z values of 2.61 and 2.40, respectively, P<0.05). Conclusions The prepared PCL negative pressure material and PBS negative pressure material have good biocompatibility, and can successfully construct the new matrix of full-thickness skin defect wounds in rats. PCL negative pressure material is better than PBS negative pressure material in general.

Clinical effects of free peroneal artery perforator flaps in repairing forefoot skin and soft tissue defect wounds assisted with three-dimensional computed tomography angiography

Wang Chengde, Wang Ai, Sun Jiling, Ma Wenguo, Wang Jianguo

2022, 38(7): 661-666. doi: 10.3760/cma.j.cn501120-20210914-00317

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Objective To investigate the clinical effects of free peroneal artery perforator flaps in repairing forefoot skin and soft tissue defect wounds assisted with three-dimensional computed tomography angiography (3D-CTA). Methods A retrospective observational study was conducted. From March 2017 to September 2019, 15 patients with skin and soft tissue defect wounds in the forefoot were treated in the Department of Burn and Plastic Surgery of Yidu Central Hospital of Weifang, including 12 males and 3 females, with age of 18-60 years. The wound area on admission was 3.0 cm×3.0 cm-9.0 cm×8.0 cm. The 3D-CTA examination before operation was performed to select the peroneal artery perforating vessels with appropriate length of vascular pedicle and good blood perfusion. According to the wound area and the perforating vessels of the peroneal artery located by 3D-CTA, the peroneal artery perforator flaps of 3.5 cm×3.5 cm-9.5 cm×8.5 cm carried with lateral sural cutaneous nerve was designed and cut, and the nerve was anastomosed with the nerve of the wound. The wound in the donor site of the flap was directly sutured or covered with medium-thickness skin graft from the thigh. The consistencies of type, diameter, and perforating position of perforating vessel of the peroneal artery detected by 3D-CTA before the operation with those of the actual measurement during operation were observed. The length of time for flap cutting and the survival of the flap after operation were recorded. During follow-up of 12 months after the operation, the patients were instructed to evaluate the foot function according to the Maryland foot function score standard, and the wound healing in the donor area and the occurrence of complications affecting the motor function of limb were observed. Data were statistically analyzed with paired sample t test. Results The types of peroneal artery perforating vessels in patients measured during the operation were septocutaneous perforator of 12 cases, musculocutaneous perforator of 2 cases, and musculomuscular septal perforator of 1 case, which were consistent with those measured by preoperative 3D-CTA. The diameter of the peroneal artery perforating vessel measured by preoperative 3D-CTA was (1.38±0.17) mm, which was close to (1.40±0.19) mm measured during the operation (t=0.30, P>0.05). The horizontal distance from the starting point of the perforating vessel to the outer edge of the shank was (42±6) mm, and the vertical distance from the starting point of the perforating vessel to the level of the lateral ankle tip was (219±14) mm measured by preoperative 3D-CTA, which were respectively close to (43±6) and (221±15) mm of intraoperative measurement (with t values of 0.46 and 0.38, respectively, P>0.05). The length of time for cutting flap was (31±6) min. All flaps survived post operation without vascular crisis. During follow-up of 12 months after the operation, the foot function was evaluated as excellent in 11 cases, good in 3 cases, and fair in 1 case, the donor site wound healed well, the scar was not noticeable with no contracture, and the motor function of joints was not affected. Conclusions Free peroneal artery perforator flap is one of the effective methods to reconstruct skin and soft tissue defect wounds in the forefoot, and the risk of surgery can be reduced when the anatomical location of the perforating vessels is confirmed by 3D-CTA.

Effects and molecular mechanism of exogenous L-carnitine on excessive endoplasmic reticulum stress-mediated hepatic pyroptosis in severely scald rats

Fan Fuxiao, Li Pengtao, Xia Zhengguo, Xie Chaoqiong, Xu Jiegou, Xu Qinglian

2022, 38(7): 667-676. doi: 10.3760/cma.j.cn501225-20220120-00010

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Objective To investigate the effects and molecular mechanism of exogenous L-carnitine on hepatic pyroptosis mediated by excessive endoplasmic reticulum stress in severely scald rats. Methods The experimental research method was adopted. According to the random number table (the same group method below), fifteen female Sprague Dawley rats aged 6-8 weeks were divided into sham-injury group, scald alone group, and scald+carnitine group (with 5 rats in each group), and full-thickness scald of 30% total body surface area were made on the back of rats in scald alone group and scald+carnitine group, and rats in scald+carnitine group were additionally given intraperitoneal injection of L-carnitine. At post injury hour (PIH) 72, The levels of aspartate aminotransferase (AST) and alanine dehydrogenase (ALT) of biochemical indicators of liver injury were detected by automatic biochemical analyzer with the sample number of 5. At PIH 72, liver tissue damage was detected by hematoxylin-eosin staining. At PIH 72, The mRNA levels of nucleotide-binding oligomerization domain-containing protein-like receptor family pyrin domain containing 3 (NLRP3), cysteine aspartic acid specific protease 1 (caspase-1), gasderminD (GSDMD), and interleukin 1β(IL-1β) in liver tissue as pyroptosis-related markers and glucose regulatory protein 78 (GRP78) and CCAAT/enhancer binding protein homologous protein (CHOP) in liver tissue as endoplasmic reticulum stress-related markers were detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction (RT-qPCR). Protein expression levels of GRP78, CHOP, NLRP3, caspase-1, caspase-1/p20, GSDMD-N, and cleaved IL-1β in liver tissue were detected by Western blotting, and the sample numbers were all 5. HepG2 cells as human liver cancer cells were divided into dimethyl sulfoxide (DMSO) group, 0.1 μmol/L tunicamycin (TM) group, 0.2 μmol/L TM group, 0.4 μmol/L TM group, and 0.8 μmol/L TM group and were treated accordingly. After 24 h of culture, cell viability was detected by cell counting kit 8, and the intervention concentration of TM was screened, and the sample number was 5. HepG2 cells were divided into DMSO group, TM alone group, and TM+carnitine group, and treated accordingly. After 24 h of culture, the protein expression levels of GRP78, CHOP, NLRP3, caspase-1, caspase-1/p20, GSDMD-N, and cleaved IL-1β in cells were detected by Western blotting, and the sample numbers were all 3. Data were statistically analyzed with one-way analysis of variance and least significant difference-t test. Results At PIH 72, the AST and ALT levels of serum in scald alone group were (640±22) and (157±8) U/L, which were significantly higher than (106±13) and (42±6) U/L in sham-injury group, respectively, with t values of -46.78 and -25.98, respectively, P<0.01. The AST and ALT levels of serum in scald+carnitine group were (519±50) and (121±10) U/L, which were significantly lower than those in scald alone group, respectively, with t values of 4.93 and 6.06, respectively, P<0.01. At PIH 72, the morphology of liver tissue of rats in sham-injury group were basically normal with no obvious inflammatory cell infiltration; compared with those in sham-injury group, the liver tissue of rats in scald alone group showed a large number of inflammatory cell infiltration and disturbed cell arrangement; compared with that in scald alone group, the liver tissue of rats in scald+carnitine group showed a small amount of inflammatory cell infiltration. At PIH 72, the mRNA expression on levels of NLRP3, caspase-1, GSDMD, and IL-1β in liver tissue of rats in scald alone group were significantly higher than those in sham-injury group (with t values of 34.42, 41.93, 30.17, and 15.68, respectively, P<0.01); the mRNA levels of NLRP3, caspase-1, GSDMD, and IL-1β in liver tissue of rats in scald+carnitine group were significantly lower than those in scald alone group (with t values of 34.40, 37.20, 19.95, and 7.88, respectively, P<0.01). At PIH 72, the protein expression levels of NLRP3, caspase-1, caspase-1/p20, GSDMD-N, and cleaved IL-1β in liver tissue of rats in scald alone group were significantly higher than those in sham-injury group (with t values of 12.28, 26.92, 5.20, 10.02, and 24.78, respectively, P<0.01); compared with those in scald alone group, the protein expression levels of NLRP3, caspase-1, caspase-1/p20, GSDMD-N, and cleaved IL-1β in liver tissue of rats in scald+carnitine group were significantly decreased (with t values of 10.99, 27.96, 12.69, 8.96, and 12.27, respectively, P<0.01). At PIH 72, the mRNA levels of GRP78 and CHOP in liver tissue of rats in scald alone group were significantly higher than those in sham-injury group (with t values of 21.00 and 16.52, respectively, P<0.01), and the mRNA levels of GRP78 and CHOP in liver tissue of rats in scald+carnitine group were significantly lower than those in scald alone group (with t values of 8.92 and 8.21, respectively, P<0.01); the protein expression levels of GRP78 and CHOP in liver tissue of rats in scald alone group were significantly higher than those in sham-injury group (with t values of 22.50 and 14.29, respectively, P<0.01), and the protein expression levels of GRP78 and CHOP in liver tissue of rats in scald+carnitine group were significantly lower than those in scald alone group (with t values of 14.29 and 5.33 respectively, P<0.01). After 24 h of culture, the cell survival rates of 0.1 μmol/L TM group, 0.2 μmol/L TM group, 0.4 μmol/L TM group, and 0.8 μmol/L TM group were significantly decreased than that in DMSO group (with t values of 4.90, 9.35, 18.64, and 25.09, respectively, P<0.01). Then 0.8 μmol/L was selected as the intervention concentration of TM. After 24 h of culture, compared with that in DMSO group, the protein expression levels of GRP78 and CHOP in cells in TM alone group were significantly increased (with t values of 10.48 and 17.67, respectively, P<0.01), and the protein expression levels of GRP78 and CHOP in TM+carnitine group were significantly lower than those in TM alone group (with t values of 8.08 and 13.23, respectively, P<0.05 or P<0.01). After 24 h of culture, compared with those in DMSO group, the protein expression levels of NLRP3 and GSDMD-N in cells in TM alone group were significantly increased (with t values of 13.44 and 27.51, respectively, P<0.01), but the protein expression levels of caspase-1, caspase-1/p20, and cleaved IL-1β in cells were not significantly changed (P>0.05); compared with that in TM alone group, the protein expression levels of NLRP3 and GSDMD-N in cells in TM+carnitine group were significantly decreased (with t values of 20.49 and 21.95, respectively, P<0.01), but the protein expression levels of caspase-1, caspase-1/p20, and cleaved IL-1β in cells were not significantly changed (P>0.05). Conclusions In severely scald rats, exogenous L-carnitine may play a protective role against liver injury by inhibiting the pathways related to excessive endoplasmic reticulum stress-mediated pyroptosis.

Clinical effects of free hallux-nail flap combined with the second toe composite tissue flap in the reconstruction of damaged thumb after electrical burns

Xing Peipeng, Mu Xinling, Xia Chengde, Shi Jijing, Xue Jidong, Yang Gaoyuan, Zhang Jian, Di Haiping

2022, 38(7): 677-682. doi: 10.3760/cma.j.cn501120-20210621-00222

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Objective To explore the clinical effects of free hallux-nail flap combined with the second toe composite tissue flap in the reconstruction of damaged thumb after electrical burns. Methods A retrospective observational study was conducted. From May 2018 to April 2021, 12 male patients with thumb destructive defects caused by electrical burns who met the inclusion criteria were admitted to Zhengzhou First People's Hospital, aged 27 to 58 years, including 10 cases with degree Ⅲ thumb defect and 2 cases with degree Ⅳ thumb defect after thorough debridement. The thumb was reconstructed with free hallux-nail flap combined with composite tissue flap of the second phalangeal bone, joint, and tendon with skin island. The donor site of hallux-nail flap was covered with artificial dermis in the first stage and performed with continuous vacuum sealing drainage, and covered with medium-thickness skin graft from the groin site in the second stage. The donor site in the second toe was filled and fixed with iliac bone strips. The survival of reconstructed thumb was observed 1 week after the reconstruction surgery, the survival of skin graft in the donor site of hallux-nail flap was observed 2 weeks after skin grafting, and the callus formation of the reconstructed thumb phalanx and the second toe of the donor foot was observed by X-ray 6 weeks after the reconstruction surgery. During the follow-up, the shape of reconstructed thumb was observed and the sensory function was evaluated; the function of reconstructed thumb was evaluated with trial standard for the evaluation of the functions of the upper limbs of the Hand Surgery Society of the Chinese Medical Association; whether the interphalangeal joints of the hallux and the second toe were stiff, the scar hyperplasia of the foot donor site, and whether the walking and standing functions of the donor feet were limited were observed. Results One week after the reconstruction surgery, all the reconstructed thumbs of the patients survived. Two weeks after skin grafting, the skin grafts in the donor site of hallux-nail flap of 11 patients survived, while the skin graft in the donor site of hallux-nail flap of 1 patient was partially necrotic, which was healed completely after 10 days' dressing change. Six weeks after the reconstruction surgery, callus formation was observed in the reconstructed thumb and the second toe of the donor foot of 10 patients, the Kirschner wires were removed; while callus formation of the reconstructed thumb was poor in 2 patients, and the Kirschner wires were removed after 2 weeks of delay. During the follow-up of 6 to 24 months, the shape of reconstructed thumb was similar to that of the healthy thumb, the discrimination distance between the two points of the reconstructed thumb was 7 to 11 mm, and the functional evaluation results were excellent in 4 cases, good in 6 cases, and fair in 2 cases. The interphalangeal joints of the hallux and the second toe of the donor foot were stiff, mild scar hyperplasia was left in the donor site of foot, and the standing and walking functions of the donor foot were not significantly limited. Conclusions The application of free hallux-nail flap combined with the second toe composite tissue flap in the reconstruction of damaged thumb after electrical burns adopts the concept of reconstruction instead of repair to close the wound. It can restore the shape and function of the damaged thumb without causing great damage to the donor foot.

A prospective randomized controlled study on the effects of compound analgesia in ultra-pulsed fractional carbon dioxide laser treatment of post-burn hypertrophic scars in children

Yang Jiao, Shi Shan, Wang Ling, Li Na, Han Juntao, Hu Dahai

2022, 38(7): 683-690. doi: 10.3760/cma.j.cn501120-20210507-00171

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Objective To investigate the effects of compound analgesia on ultra-pulsed fractional carbon dioxide laser (UFCL) treatment of post-burn hypertrophic s in children. Methods A prospective randomized controlled study was conducted. From April 2018 to March 2020, 169 pediatric patients with post-burn hypertrophic s admitted to the First Affiliated Hospital of Air Force Medical University were randomly divided into general anesthesia alone group (39 cases, 19 males and 20 females, aged 35 (21, 48) months), general anesthesia+lidocaine group (41 cases, 23 males and 18 females, aged 42 (22, 68) months), general anesthesia+ibuprofen suppository group (41 cases, 25 males and 16 females, aged 38 (26, 52) months), and three-drug combination group with general anesthesia + lidocaine+ibuprofen suppository (48 cases, 25 males and 23 females, aged 42 (25, 60) months), and the pediatric patients in each group were treated with corresponding analgesic regimens when UFCL was used to treat s, and the pediatric patients were given comprehensive care throughout the treatment process. The pain degree of pediatric patients scar was evaluated by facial expression,legs,activity,cry,and consolability (FLACC) of children's pain behavior scale at 0 (immediately), 1, 2, and 4 h after awakening from the first anesthesia, respectively. At 4 h after awakening from the first anesthesia of postoperative pain assessment, the self-made analgesia satisfaction questionnaire was used to evaluate the satisfaction for the analgesic effect of the pediatric patients or their families, and the satisfaction rate was calculated. Within 2 h after the first operation, the occurrences of adverse reactions of the pediatric patients, such as nausea and vomiting, headache, dizziness, drowsiness, etc, were observed and recorded. Before the first treatment and 1 month after the last treatment, the Vancouver scar scale (VSS) was used to evaluate the pediatric patients scar, and the difference value between the two was calculated. Data were statistically analyzed with least significant difference test, Kruskal-Wallis H test, chi-square test and Fisher's exact probability test. Results At 0 h after awakening from the first anesthesia, the FLACC scores of pediatric patients in general anesthesia+lidocaine group, general anesthesia+ibuprofen suppository group and three-drug combination group were significantly lower than those in general anesthesia alone group (P<0.01). The FLACC scores of the pediatric patients in anesthesia+ibuprofen suppository group and three-drug combination group were significantly lower than that in general anesthesia+lidocaine group (P<0.01), and the FLACC score of the pediatric patients in three-drug combination group was significantly lower than that in general anesthesia+ibuprofen suppository group (P<0.01). At 1 and 2 h after awakening from the first anesthesia, the FLACC scores of pediatric patients in general anesthesia+ibuprofen suppository group and three-drug combination group were both significantly lower than those in general anesthesia alone group and general anesthesia+lidocaine group (P<0.01), and the FLACC score of the pediatric patients in three-drug combination group was significantly lower than that in general anesthesia+ibuprofen suppository group (P<0.01). At 4 h after awakening from the first anesthesia, the FLACC scores of the pediatric patients in general anesthesia+ibuprofen suppository group and three-drug combination group were significantly lower than those in general anesthesia alone group and general anesthesia+lidocaine group (P<0.01). At 4 h after awakening from the first anesthesia, the satisfactions rate with the analgesic effect in the four groups of pediatric patients or their families were 79.49% (31/39), 85.37% (35/41), 87.80% (36/41), and 97.92% (47/48), respectively. The satisfaction rate of the pediatric patients in three-drug combination group was significantly higher than those in general anesthesia alone group, general anesthesia+lidocaine group, general anesthesia+ibuprofen suppository group. Within 2 h after the first operation, there was no significant difference in the overall comparison of adverse reactions such as nausea and vomiting, headache, dizziness, and drowsiness of pediatric patients among the 4 groups (P>0.05). The VSS scores of pediatric patients before the first treatment, 1 month after the last treatment, and and the difference value between the two in the 4 groups were not significantly different (P>0.05). Conclusions Three-drug combination for analgesia has a good effect in the treatment of hypertrophic scars after burn in pediatric patients with UFCL. Pediatric patients or their families are highly satisfied with the effect, and the treatment effect and incidence of adverse reactions are similar to other analgesic regimens, so it is recommended to be promoted in clinical practice.

Research advances on biomaterials for the delivery of growth factors to regulate wound repair

Weng Tingting, Cai Chenghao, Han Chunmao, Wang Xingang

2022, 38(7): 691-696. doi: 10.3760/cma.j.cn501225-20220430-00166

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Wound repair is a highly coordinated and mutually regulated complex process involving various kinds of cells, extracellular matrices and cytokines. A variety of growth factors play an important regulatory role in wound healing, and it is critical to achieve effective delivery and sustained function of growth factors. In recent years, the application of biomaterials in tissue engineering has shown great potential, and the effective delivery of growth factors by biomaterials has attracted increasing attention. Based on this, this paper introduces the mechanism of related growth factors in the process of wound healing, focusing on the recent progress of biomaterial delivery of growth factors to accelerate wound healing, in order to provide new enlightenment for clinical wound treatment.

Research advances on the application of carbon dots in wound treatment

Wang Peng, Wang Guangyi, Ji Shizhao, Ma Jianming, Tang Tao

2022, 38(7): 697-704. doi: 10.3760/cma.j.cn501120-20210709-00242

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Chronic and infectious wound healing has always been an issue of concern in clinical and scientific research, in which bacterial infection and oxidative damage are the key factors hindering wound healing. Carbon dots, as a new material, has attracted much attention because of its unique physical and chemical properties and good biological safety. In recent years, the researches on the antibacterial property, antioxidant, and photoluminescence properties of carbon dots are more and more extensive and carbon dots have great potential in the treatment of chronic and infectious wounds. This paper reviews the research progress of carbon dots in three aspects: antibacterial, anti-oxidation and monitoring of wound infection are reviewed, and further discusses its specific mechanism, potential research direction, and application prospect.

2022 Vol. 38, No. 7

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