Nevin Manimala

Zhonghua Shao Shang Za Zhi. 2023 Jun 20;39(6):565-572. doi: 10.3760/cma.j.cn501225-20220903-00377.

ABSTRACT

Objective: To explore the effects of tensile force on vascular lumen formation in three-dimensional printed tissue. Methods: The experimental research method was used. Human umbilical vein endothelial cells (HUVECs) were extracted from discarded umbilical cord tissue of 3 healthy women (aged 22 to 35 years) who gave birth in the Department of Gynaecology and Obstetrics of Suzhou Ruihua Orthopaedic Hospital from September 2020 to May 2021. Human skin fibroblasts (HSFs) were extracted from discarded normal skin tissue of 10 male patients (aged 20 to 45 years) who underwent wound repair in the Department of Hand Surgery of Suzhou Ruihua Orthopaedic Hospital from September 2020 to September 2022. After identification of the two kinds of cells, the 4^th to 6^th passage of cells were taken for the follow-up experiments. HUVECs and HSFs were used as seed cells, and polycaprolactone, gelatin, hyaluronic acid, and fibrin were used as scaffold materials, and the three-dimensional printed vascularized tissue was created by three-dimensional bioprinting technology. The printed tissue with polycaprolactone scaffold of 6 and 10 mm spacing, and without polycaprolactone scaffold were set as 6 mm spacing polycaprolactone group, 10 mm spacing polycaprolactone group, and non-polycaprolactone group, respectively. After 4 days of culture, the printed tissue in 10 mm spacing polycaprolactone group was selected to detect the cell survival by cell viability detection kit, and the cell survival rate was calculated. After 14 days of culture, the printed tissue in three groups were taken, and the shape change of tissue was observed by naked eyes; immunofluorescence staining was performed to observe the arrangement of filamentous actin, and lumen diameter, total length, and number of branches of vessel in the tissue. The tissue with micro-spring structure in the above-mentioned three groups was designed, printed, and cultured for 9 days, and the tensile force applied in the printed tissue was measured according to the force-displacement curve. The number of samples was all 3 in the above experiments. Data were statistically analyzed with one-way analysis of variance and Tukey test. Results: After 4 days of culture, the cell survival rate in printed tissue in 10 mm spacing polycaprolactone group was (91.3±2.2)%. After 14 days of culture, the shape change of printed tissue in non-polycaprolactone group was not obvious, while the shape changes of printed tissue in 6 mm spacing polycaprolactone group and 10 mm spacing polycaprolactone group were obvious. After 14 days of culture, the arrangement of filamentous actin in the printed tissue in non-polycaprolactone group had no specific direction, while the arrangement of filamentous actin in the printed tissue in 6 mm spacing polycaprolactone group and 10 mm spacing polycaprolactone group had a specific direction. After 14 days of culture, The vascular lumen diameters of the printed tissue in 6 mm spacing polycaprolactone group and 10 mm spacing polycaprolactone group were (6.0±1.3) and (10.8±1.3) μm, respectively, which were significantly larger than 0 μm in non-polycaprolactone group (P<0.05), and the vascular lumen diameter of printed tissue in 10 mm spacing polycaprolactone group was significantly larger than that in 6 mm spacing polycaprolactone group (P<0.05); the total length and number of branches of blood vessel in the printed tissue in 6 mm spacing polycaprolactone group and 10 mm spacing polycaprolactone group were significantly shorter or less than those in non-polycaprolactone group (P<0.05), and the total length and number of branches of blood vessel in the printed tissue in 10 mm spacing polycaprolactone group were significantly shorter or less than those in 6 mm spacing polycaprolactone group. After 9 days of culture, the tensile forces applied in the printed tissue in 6 mm spacing polycaprolactone group and 10 mm spacing polycaprolactone group were (2 340±59) and (4 284±538) μN, respectively, which were significantly higher than 0 μN in non-polycaprolactone group (P<0.05), and the tensile force applied in the printed tissue in 10 mm spacing polycaprolactone group was significantly higher than that in 6 mm spacing polycaprolactone group (P<0.05). Conclusions: The three-dimensional printed scaffold structure can exert different tensile force in the printed tissue, and the vascular lumen diameter of the printed tissue can be regulated by adjusting the tensile force.

PMID:37805773 | DOI:10.3760/cma.j.cn501225-20220903-00377

Zhonghua Shao Shang Za Zhi. 2023 Jun 20;39(6):558-564. doi: 10.3760/cma.j.cn501225-20220806-00336.

ABSTRACT

Objective: To explore the epidemiological characteristics and risk factors of sepsis development and death in patients with extremely severe burns. Methods: A retrospective case series study was conducted. From January 2017 to December 2021, 135 patients with extremely severe burns who met the inclusion criteria were admitted to the Department of Burn and Wound Repair of the Second Affiliated Hospital of Zhejiang University School of Medicine, including 100 males and 35 females, aged 18-84 years. The incidence and diagnosis time of sepsis, the rate of positive microbial culture of blood samples (hereinafter referred to as positive blood culture), and the mortality rate of all patients, as well as the incidence of sepsis and the pathogen of infection in patients with positive blood culture were recorded (statistically analyzed with chi-square test or Fisher’s exact probability test). According to the occurrence of sepsis, all patients were divided into sepsis group (58 cases) and non-sepsis group (77 cases), and the gender, age, body mass index, history of hypertension, history of diabetes, combination of inhalation injury, burn site, burn type, total burn area, and combined injury of patients were compared between the two groups. According to the outcome, all patients were divided into death group (37 cases) and survival group (98 cases), and the aforementioned data grouped according to sepsis as well as the stability of shock period and the combination of sepsis of patients were compared between the two groups. The aforementioned data between two groups were statistically analyzed with univariate analysis of independent sample t test, Wilcoxon rank-sum test, Mann-Whitney U test, chi-square test, or Fisher’s exact probability test. Factors with P<0.1 were selected for multivariate logistic regression analysis to screen independent risk factors of sepsis and death in patients with extremely severe burns. Results: Among all patients, the incidence of sepsis was 42.96% (58/135), the diagnosis time of sepsis was 14 (7, 24) d after injury, the positive blood culture rate was 62.22% (84/135), and the mortality rate was 27.41% (37/135). The incidence of sepsis of patients with positive blood culture was 69.05% (58/84). The top 5 pathogenic bacteria in the detection rate of septic patients with positive blood culture were Acinetobacter baumannii, Klebsiella pneumoniae, Pseudomonas aeruginosa, Staphylococcus aureus, and Enterobacter cloacae, ranking from high to low, and the proportion of Acinetobacter baumannii infected was significantly higher than that of non-septic patients with positive blood culture (χ²=7.49, P<0.05). Compared with those in non-sepsis group, the proportion of combination of inhalation injury, the proportion of perineal burns, and the total burn area of patients in sepsis group increased significantly (with χ² values of 11.08 and 17.47, respectively, Z=5.68, P<0.05), while the other indicators did not change significantly (P>0.05). Multivariate logistic regression analysis showed that combination of inhalation injury, total burn area ≥80% total body surface area (TBSA), and perineal burns were independent risk factors for patients with extremely severe burns developing sepsis (with odds ratios of 3.15, 7.24, and 3.24, respectively, with 95% confidence intervals of 1.07 to 9.29, 1.79 to 29.34, and 1.21 to 8.68, respectively, P<0.05). Compared with those in survival group, the proportion of combination of inhalation injury, the proportion of perineal burns, and the proportion of combination of sepsis (with χ² values of 6.55, 11.64, and 22.26, respectively, P values all <0.05), total burn area (Z=4.25, P<0.05), and proportion of instability of shock period (P<0.05) of patients in death group all increased significantly, while the other indicators did not change significantly (P>0.05). Multivariate logistic regression analysis showed that the instability of shock period and combination of sepsis were independent risk factors for death of patients with extremely severe burns (with odds ratios of 4.87 and 3.45, respectively, with 95% confidence intervals of 1.21 to 19.57 and 1.28 to 9.33, respectively, P<0.05). Conclusions: Patients with extremely severe burns have a high incidence of sepsis and a high mortality rate. The peak period of sepsis onset is 2 weeks after injury, with Acinetobacter baumannii as the most prominent infectious pathogen. Combination of inhalation injury, total burn area ≥80% TBSA, and perineal burns are independent risk factors for extremely severe burn patients complicated with sepsis, and combination of sepsis and instability of shock period are independent risk factors for death of patients with extremely severe burns.

PMID:37805772 | DOI:10.3760/cma.j.cn501225-20220806-00336

Zhonghua Shao Shang Za Zhi. 2023 Jun 20;39(6):552-557. doi: 10.3760/cma.j.cn501225-20220714-00294.

ABSTRACT

Objective: To compare the curative effects of butterfly-shaped flap based on the dorsal branch of digital artery (hereinafter referred to as butterfly-shaped flap) and propeller flap based on the dorsal branch of digital artery (hereinafter referred to as propeller flap) in repairing the wound in volar aspect of finger. Methods: A retrospective cohort study was conducted. From August 2018 to April 2022, 16 patients with finger palmar wounds admitted to Ruijin Hospital of Shanghai Jiao Tong University School of Medicine and 7 patients with finger palmar wounds admitted to General Hospital of PLA Central Theater Command met the inclusion criteria, including 14 males and 9 females, aged 25 to 64 years. After debridement or resection of skin benign tumor, the wounds ranged from 0.5 cm×0.5 cm to 1.5 cm×1.5 cm. According to the different rotation axes of flap pedicle during wound repair, the patients were divided into butterfly-shaped flap group (8 cases) and propeller flap group (15 cases), and their wounds were repaired by butterfly-shaped flap (with area of 0.5 cm×0.5 cm-1.5 cm×1.3 cm) or propeller flap (with area of 0.7 cm×0.5 cm-1.5 cm×1.5 cm) , respectively. In propeller flap group, wounds in the donor sites were repaired by full-thickness skin grafts taken from the palms of wrists or the groin. The surgical time, postoperative complications, flap survival, and wound healing time of patients in the two groups were recorded. Data were statistically analyzed with independent sample t test, Mann Whitney U test, or Fisher’s exact probability test. Results: The surgical time and postoperative wound healing time of patients in butterfly-shaped flap group ((43±9) min and (13.1±0.8) d, respectively) were both significantly shorter than those in propeller flap group ((87±16) min and (16.7±4.6) d, respectively, with t values of -7.03 and -2.86, respectively, P<0.05). The postoperative flap survival and complications of patients between the two groups were both similar (P>0.05). Conclusions: For repairing the wound in volar aspect of finger, the butterfly-shaped flap has more advantages in comparison with the traditional propeller flap. The butterfly-shaped flap has a short surgical time and fast postoperative recovery, which is worthy of clinical promotion.

PMID:37805771 | DOI:10.3760/cma.j.cn501225-20220714-00294

Zhonghua Shao Shang Za Zhi. 2023 Jun 20;39(6):518-526. doi: 10.3760/cma.j.cn501225-20230421-00137.

ABSTRACT

Objective: To explore the biological role and clinical significance of ubiquitin-specific protease 7 (USP7) in the carcinogenesis of scar ulcer. Methods: A retrospective observational study combined with bioinformatics analysis was used. The RNA expression profile data of USP7 in tumor and/or its corresponding paracancular normal tissue were obtained from The Cancer Genome Atlas (TCGA) database and the Gene Expression Omnibus database, and the RNA sequencing data were transformed by log₂. The variations of USP7 gene were analyzed by cBioPortal database. The USP7 mRNA expression in tumor and adjacent normal tissue in TCGA database were obtained by using the “Gene_DE” module in TIMER 2.0 database. The survival rates of patients with high and low USP7 expression in cutaneous melanoma (SKCM), cervical squamous cell carcinoma (CESC), lung squamous cell carcinoma (LUSC), and head and neck squamous cell carcinoma (HNSC) were analyzed using the Gene Expression Profile Interactive Analysis 2 (GEPIA2) database, and the Kaplan-Meier survival curves were drawn. Sangerbox database was used to analyze the correlation of USP7 expression in pan-cancer with microsatellite instability (MSI) or tumor mutation burden (TMB) pan-cancer. Through the “correlation analysis” module in the GEPIA2 database, the correlation of USP7 expression in pan-cancer with the expression levels of five DNA mismatch repair genes (MLH1, MSH2, MSH6, PMS2, and EPCAM) and three essential DNA methyltransferases (DNMT)–DNMT1, DNMT3A, and DNMT3B were evaluated. The USP7 expression in CESC, HNSC, LUSC, and SKCM and its correlation with infiltration of immune cells (B cells, CD4⁺ T cells, CD8⁺ T cells, neutrophils, macrophages, and dendritic cells) were analyzed by the “Immune-Gene” module in TIMER 2.0 database. The “Similar Genes Detection” module of GEPIA2 database was used to obtain the top 100 protein sets with similar expression patterns to USP7. Intersection analysis was performed between the aforementioned protein sets and the top 50 protein sets that were directly physically bound to USP7 obtained by using the STRING database. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analysis were performed for the two protein sets mentioned above using the DAVID database. The samples of normal skin, hypertrophic scar, scar ulcer, and scar carcinoma with corresponding clinicopathologic features were collected from the Department of Pathology of Tongren Hospital of Wuhan University & Wuhan Third Hospital from October 2018 to October 2022, and the USP7 expression in tissue was detected by immunohistochemical method, with the number of samples of 6. Data were statistically analyzed with Log-rank test, one-way analysis of variance, and Bonferroni test. Results: In pan-cancer, the main gene variations of USP7 were mutation and amplification, and the top 3 tumors with the highest variation frequency (>6%) were bladder urothelial carcinoma, SKCM, and endometrial carcinoma. The main mutation of USP7 gene in pan-cancer was missense mutation. In SKCM with the highest mutation frequency, the main type of mutation was missense mutation in USP7_ICP0_bdg domain. USP7 mRNA expression in breast invasive carcinoma, bile duct carcinoma, colon carcinoma, esophageal carcinoma, HNSC, renal chromophobe cell carcinoma, hepatocellular carcinoma, lung adenocarcinoma, LUSC, prostate carcinoma, and gastric carcinoma was significantly higher than that in corresponding paracancer normal tissue (P<0.05). USP7 mRNA expression in glioblastoma multiforme, renal clear cell carcinoma, renal papillary cell carcinoma, and thyroid carcinoma was significantly lower than that in corresponding paracancular normal tissue (P<0.05). In addition, USP7 mRNA expression in SKCM metastases was much higher than that in primary tumor tissue (P<0.05). Survival curves showed no significant difference in survival rate between patients with high USP7 expression and patients with low USP7 expression in CESC, HNSC, LUSC, and SKCM (Log-rank P>0.05, with hazard ratios of 1.00, 0.99, 1.00, and 1.30, respectively). USP7 expression in colon cancer, colorectal cancer, thymic cancer, and thyroid cancer was negatively correlated with TMB (with Pearson correlation coefficients of -0.26, -0.19, -0.19, and 0.11, respectively, P<0.05). USP7 expression in glioma, CESC, lung adenocarcinoma, mixed renal carcinoma, and LUSC was positively correlated with MSI expression (with Pearson correlation coefficients of 0.22, 0.14, 0.15, 0.08, and 0.14, respectively, P<0.05), and USP7 expression in colon cancer, colorectal cancer, invasive breast cancer, prostate cancer, HNSC, thyroid cancer, and diffuse large B-cell lymphoma were significantly negatively correlated with MSI expression (with Pearson correlation coefficients of -0.31, -0.27, -0.13, -0.19, -0.16, -0.18, and -0.53, respectively, P<0.05). The expression of USP7 in CESC was positively correlated with that of both MSH2 and MSH6 (with Spearman correlation coefficients of 0.51 and 0.44, respectively, P<0.05), and the expression of USP7 in HNSC was positively correlated with the expression of EPCAM, MLH1, MSH2, MSH6, and PMS2 (with Spearman correlation coefficients of 0.39, 0.14, 0.49, 0.54, and 0.41, respectively, P<0.05), and the expression of USP7 in LUSC was positively correlated with the expression of EPCAM, MSH2, MSH6, and PMS2 (with Spearman correlation coefficients of 0.20, 0.36, 0.40, and 0.34, respectively, P<0.05), and the expression of USP7 in SKCM was positively correlated with the expression of EPCAM, MLH1, MSH2, MSH6, and PMS2 (with Spearman correlation coefficients of 0.11, 0.33, 0.42, 0.55, and 0.34, respectively, P<0.05). The expression of USP7 in CESC, HNSC, LUSC, and SKCM was significantly positively correlated with the expression of DNMT1, DNMT3A, and DNMT3B (with Spearman correlation coefficients of 0.42, 0.34, 0.22, 0.45, 0.52, 0.22, 0.36, 0.36, 0.22, 0.38, 0.46, and 0.21, respectively, P<0.05). The expression of USP7 in CESC, HNSC, LUSC, and SKCM was positively correlated with CD4⁺ T cell infiltration (with Partial correlation coefficients of 0.14, 0.22, 0.13, and 0.16, respectively, P<0.05). Being similar to the pattern of USP7 expression and ranked among top 100 protein sets, the top 5 proteins were C16orf72, BCLAF1, UBN, GSPT1, ERI2 (with Spearman correlation coefficients of 0.83, 0.74, 0.73, and 0.72, respectively, all P values<0.05). The top 50 protein sets that directly physically bind to USP7 overlapped with the aforementioned protein set by only one protein, thyroid hormone receptor interaction factor 12. KEGG enrichment analysis showed that USP7 related genes were involved in cell cycle, spliceosome, cell senescence, and p53 signal pathway. GO enrichment analysis showed that USP7 related genes were involved in transcriptional regulation, protein ubiquitination, DNA repair, and cytoplasmic pattern recognition receptor signal pathways. Analysis of clinical samples showed that USP7 expression was significantly higher in hypertrophic scars (0.35±0.05), scar ulcers (0.43±0.04), and scar cancers (0.61±0.03) than in normal skin (0.18±0.04), P<0.05. Conclusions: USP7 may be a clinical biomarker for the progression of cicatricial ulcer cancer.

PMID:37805766 | DOI:10.3760/cma.j.cn501225-20230421-00137

Zhonghua Shao Shang Za Zhi. 2023 Jun 20;39(6):512-517. doi: 10.3760/cma.j.cn501225-20230116-00018.

ABSTRACT

Objective: To compare the efficacy and safety of 2 940 nm fractional erbium laser combined with fractional micro-plasma radiofrequency (FMR) therapy and 2 940 nm fractional erbium laser in the treatment of atrophic acne scars. Methods: A prospective randomized controlled research was conducted. A total of 100 atrophic acne scar patients (38 males and 62 females, aged 18-37 years) who were treated in the Scar Laser Clinic of the Department of Plastic and Reconstructive Surgery of Shanghai Ninth People’s Hospital of Shanghai Jiao Tong University School of Medicine from March 2018 to March 2021 and conformed to the inclusion criteria were recruited. The patients were randomly divided into erbium laser+FMR group and erbium laser alone group, with 50 cases in each group. The facial acne scars of patients in erbium laser alone group were treated with 2 940 nm fractional erbium laser, while the facial acne scars of patients in erbium laser+FMR group were treated with erbium laser as above, besides, the scars of U and M types were treated with FMR, once every 3 months for a total of 3 times. Before the first treatment and 3 months after each treatment, the Echelle D’Assessment Clinique des Cicatrices D’Acne (ECCA) was used to score the scar. The occurrence of adverse reaction during the treatment process was observed and recorded, and the incidence was calculated. Three months after the last treatment, the 5-level classification method was used to evaluate the satisfaction of patients with the treatment effect, and the satisfaction rate was calculated. Data were statistically analyzed with independent sample t test and chi-square test. Results: A total of 89 patients completed the study, including 46 patients in erbium laser+FMR group (19 males and 27 females, aged (26±5) years) and 43 patients in erbium laser alone group (15 males and 28 females, aged (27±6) years). The ECCA scores before the first treatment and 3 months after the first treatment of patients were similar between the two groups (P>0.05). The ECCA scores of patients in erbium laser+FMR group at 3 months after the second and third treatment were 72±23 and 61±18, respectively, which were significantly lower than 85±26 and 76±25 in erbium laser alone group (with t values of -2.45 and -3.26, respectively, P<0.05). During the treatment process, the incidence of adverse reaction of patients in erbium laser+FMR group and erbium laser alone group were 23.91% (11/46) and 16.28% (7/43), respectively, and there was no statistically significant difference between the two groups (P>0.05). The satisfaction rate of patients in erbium laser+FMR group was 78.26% (36/46) at 3 months after the last treatment, which was significantly higher than 53.49% (23/43) in erbium laser alone group (χ²=6.10, P<0.05). Conclusions: The 2 940 nm fractional erbium laser combined with FMR is superior to 2 940 nm fractional erbium laser alone in the treatment of facial atrophic acne scars, achieving significantly higher efficacy without significantly increasing the incidence of adverse reaction, and patients are more satisfied with the efficacy. It can be used as a recommended therapy in clinical practice.

PMID:37805765 | DOI:10.3760/cma.j.cn501225-20230116-00018

Zhonghua Shao Shang Za Zhi. 2023 May 20;39(5):465-471. doi: 10.3760/cma.j.cn501225-20220524-00200.

ABSTRACT

Objective: To explore the application effects of mind mapping combined with scenario simulation training on the ability training of junior nurses in hospital transfer of patients with critical burns and trauma. Methods: A prospective randomized controlled study was conducted. From December 2019 to December 2020, 55 female junior nurses from the Institute of Burn Research of the First Affiliated Hospital of Army Medical University (the Third Military Medical University) who met the inclusion criteria were enrolled in this study and divided into routine group (27 nurses, aged (24.0±0.9) years) and combined group (28 nurses, aged (24.2±0.8) years), according to the random number table. The nurses in routine group were trained with hospital transfer of patients with critical burns and trauma by theory combined with operational skill, and the nurses in combined group were trained with hospital transfer of patients with critical burns and trauma by mind mapping combined with scenario simulation training. Before and after the training, the self-made theoretical examination papers and skill assessment items were used for the examination and assessment to nurses, and their scores were calculated and compared. The self-made emergency ability scoring system was used to evaluate the emergency disposal ability of nurses from five dimensions, including team cooperation ability, emergency response ability, operative technique ability, specialized business ability, and nurse-patient communication ability, and their scores were calculated and compared. The non-standard implementation rates of transfer nursing measures, such as incomplete preparation of goods, poor communication effect of patients, inadequate pipeline nursing, unclear handover, and inadequate final treatment, were calculated and compared in the process of transporting highly simulated human (hereinafter referred to as simulated human) by nurses before and after training; and the rate of disease change and successful rate of transport of simulated human were calculated and compared after training. After assessment, self-made satisfaction questionnaire was used to compare nurses’ satisfaction with the training mode, content, and effects. Data were statistically analyzed with independent sample t test, Pearson chi-square test, or Yates corrected chi-square test. Results: Fifty-five enrolled nurses were fully involved in the training, examination, assessment, and questionnaire filling. Before training, there were no statistically significant differences in theoretical examination and skill assessment scores between the 2 groups (P>0.05); After training, the theoretical examination and skill assessment scores of nurses in combined group were significantly higher than those in routine group (with t values of -3.89 and -4.24, respectively, P<0.05). Before training, there were no statistically significant differences in the scores of each item of emergency disposal ability between the 2 groups (P>0.05); after training, the scores in terms of team cooperation ability, emergency response ability, operative technique ability, specialized business ability, and nurse-patient communication ability of nurses in combined group were significantly higher than those in routine group (with t values of -6.49, -6.44, -2.21, -2.85, and -2.34, respectively, P<0.05). Before training, there were no statistically significant differences in the non-standard implementation rates of transfer nursing measures of nurses between the 2 groups (P>0.05); after training, the non-standard rates of incomplete preparation of goods, unclear handover, and inadequate final treatment of nurses in combined group were significantly lower than those in routine group (with t values of 3.87, 5.89, and 5.28, respectively, P<0.05). After training, the rate of disease change of simulated human of nurses in combined group was 7.14% (2/28), which was significantly lower than 33.33% (9/27) in routine group (χ²=5.89, P<0.05); the successful rate of transport was 96.43% (27/28), which was significantly higher than 74.07% (20/27) in routine group (χ²=3.87, P<0.05). After assessment, the total score of training satisfaction and scores of satisfaction with training mode and training effect of nurses in combined group were significantly higher than those in routine group (with t values of 5.22, 4.67, and 10.71, respectively, P<0.05). There was no statistically significant difference in the satisfaction score on training content between the two groups (P>0.05). Conclusions: Evidence-based mind mapping combined with scenario simulation training significantly improves the nursing skills and emergency handling capabilities of junior nurses in transferring patients with critical burns and trauma.

PMID:37805756 | DOI:10.3760/cma.j.cn501225-20220524-00200

Zhonghua Shao Shang Za Zhi. 2023 May 20;39(5):456-464. doi: 10.3760/cma.j.cn501225-20220408-00130.

ABSTRACT

Objective: To explore the effects and mechanism of annexin A1 (ANXA1)-overexpressing human adipose-derived mesenchymal stem cells (AMSCs) in the treatment of mice with acute respiratory distress syndrome (ARDS). Methods: The experimental study method was adopted. After the adult AMSCs were identified by flow cytometry, the 3^rd passage cells were selected for the follow-up experiments. According to the random number table (the same grouping method below), the cells were divided into ANXA1-overexpressing group transfected with plasmid containing RNA sequences of ANXA1 gene and no-load control group transfected with the corresponding no-load plasmid. The other cells were divided into ANXA1-knockdown group transfected with plasmid containing small interfering RNA sequences of ANXA1 gene and no-load control group transfected with the corresponding no-load plasmid. At post transfection hour (PTH) 72, the fluorescence expression was observed under a fluorescence microscope imaging system, and the protein and mRNA expressions of ANXA1 were detected by Western blotting and real-time fluorescence quantitative reverse transcription polymerase chain reaction respectively (with the sample numbers being 3). Fifty male C57BL/6J mice aged 6-8 weeks were divided into sham injury group, ARDS alone group, normal cell group, ANXA1-overexpressing group, and ANXA1-knockdown group, with 10 mice in each group. Mice in the last 4 groups were treated with endotoxin/lipopolysaccharide to make ARDS lung injury model, and mice in sham injury group were simulated to cause false injury. Immediately after injury, mice in sham injury group and ARDS alone group were injected with normal saline through the tail vein, while mice in normal cell group, ANXA1-overexpressing group, and ANXA1-knockdown group were injected with normal AMSCs, ANXA1-overexpressing AMSCs, and ANXA1-knockdown AMSCs, correspondingly. At post injection hour (PIH) 24, 5 mice in each group were selected, the Evans blue staining was performed to observe the gross staining of the right lung tissue, and the absorbance value of bronchoalveolar lavage fluid (BALF) supernatant of left lung was detected by microplate reader to evaluate the pulmonary vascular permeability. Three days after injection, the remaining 5 mice in each group were taken, the right lung tissue was collected for hematoxylin-eosin staining to observe the pathological changes and immunohistochemical staining to observe the CD11b and F4/80 positive macrophages, and the levels of tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), and IL-1β in BALF supernatant of left lung were determined by enzyme-linked immunosorbent assay. Data were statistically analyzed with paired sample t test, one-way analysis of variance, and least significant difference test. Results: At PTH 72, AMSCs in both ANXA1-overexpressing group and ANXA1-knockdown group expressed higher fluorescence intensity than AMSCs in corresponding no-load control group, respectively. At PTH 72, compared with those in corresponding no-load control group, the protein and mRNA expressions of ANXA1 in ANXA1-overexpressing group were significantly increased (wth t values of 249.80 and 6.56, respectively, P<0.05), while the protein and mRNA expressions of ANXA1 in ANXA1-knockdown group were significantly decreased (wth t values of 176.50 and 18.18, respectively, P<0.05). At PIH 24, compared with those in sham injury group (with the absorbance value of BALF supernatant being 0.041±0.009), the lung tissue of mice in ARDS alone group was obviously blue-stained and the absorbance value of BALF supernatant (0.126±0.022) was significantly increased (P<0.05). Compared with those in ARDS alone group, the degree of blue-staining in lung tissue of mice was significantly reduced in normal cell group or ANXA1-overexpressing group, and the absorbance values of BALF supernatant (0.095±0.020 and 0.069±0.015) were significantly decreased (P<0.05), but the degree of blue-staining in lung tissue and the absorbance value of BALF supernatant (0.109±0.016, P>0.05) of mice in ANXA1-knockdown group had no significant change. Compared with that in normal cell group, the absorbance value of BALF supernatant of mice in ANXA1-overexpressing group was significantly decreased (P<0.05). Three days after injection, the lung tissue structure of mice in ARDS alone group was significantly damaged compared with that in sham injury group. Compared with those in ARDS alone group, hemorrhage, infiltration of inflammatory cells, alveolar collapse, and interstitial widening in the lung tissue of mice were significantly alleviated in normal cell group and ANXA1-overexpressing group, while no significant improvement of above-mentioned lung tissue manifestation was observed in ANXA1-knockdown group. Three days after injection, the numbers of CD11b and F4/80 positive macrophages in the lung tissue of mice in ARDS alone group were significantly increased compared with those in sham injury group. Compared with those in ARDS alone group, the numbers of CD11b and F4/80 positive macrophages in lung tissue of mice in normal cell group, ANXA1-overexpressing group, and ANXA1-knockdown group reduced, with the most significant reduction in ANXA1-overexpressing group. Three days after injection, compared with those in sham injury group, the levels of TNF-α, IL-6, and IL-1β in BALF supernatant of mice in ARDS alone group were significantly increased (P<0.05). Compared with those in ARDS alone group, the levels of TNF-α, IL-6, and IL-1β in BALF supernatant of mice in normal cell group and ANXA1-overexpressing group, as well as the level of IL-1β in BALF supernatant of mice in ANXA1-knockdown group were significantly decreased (P<0.05). Compared with that in normal cell group, the level of TNF-α in BALF supernatant of mice was significantly decreased in ANXA1-overexpressing group (P<0.05) but significantly increased in ANXA1-knockdown group (P<0.05). Conclusions: Overexpression of ANXA1 can optimize the efficacy of AMSCs in treating ARDS and enhance the effects of these cells in inhibiting inflammatory response and improving pulmonary vascular permeability, thereby alleviating lung injury of mice with ARDS.

PMID:37805755 | DOI:10.3760/cma.j.cn501225-20220408-00130

Zhonghua Shao Shang Za Zhi. 2023 May 20;39(5):443-449. doi: 10.3760/cma.j.cn501225-20220608-00228.

ABSTRACT

Objective: To investigate the influence of muscle energy technology (MET) combined with Maitland joint mobilization surgery on the elbow joint flexion function in patients with deep burn of elbow joint. Methods: A retrospective controlled clinical trial was conducted. From January 2020 to January 2022, 53 patients with elbow joint flexion dysfunction after deep burns who met the inclusion criteria were treated in Tongren Hospital of Wuhan University & Wuhan Third Hospital, including 32 males and 21 females, aged (37±12) years. According to the treatment method used, the patients were divided into conventional treatment alone group (15 cases), conventional treatment+joint mobilization surgery group (18 cases), and conventional treatment+joint mobilization surgery+MET group (20 cases). Before treatment and 2 months after treatment, the patient’s elbow joint range of motion was measured using a protractor, the Mayo elbow joint function score was used to evaluate elbow joint function, a portable muscle strength tester was used to measure elbow extensor muscle strength, and visual analogue scale was used to evaluate pain degree. Data were statistically analyzed with one-way analysis of variance, least significant difference test, paired sample t test, Kruskal-Wallis H test, Wilcoxon signed rank-sum test, chi-square test, Fisher’s exact probability test, and Bonferroni correction. Results: After two months of treatment, the elbow joint range of motion and elbow joint function scores of patients in conventional treatment+joint mobilization surgery group and conventional treatment+joint mobilization surgery+MET group ((103±12)° and 60 (50, 66), (131±14)° and 73 (65, 80)) were significantly larger and higher than those in conventional treatment alone group ((77±15)° and 45 (35, 50), P values all <0.05), respectively. The elbow joint range of motion and elbow joint function scores of patients in conventional treatment+joint mobilization surgery+MET group were significantly larger and higher than those in conventional treatment+joint mobilization surgery group (P values all <0.05), respectively. After two months of treatment, the elbow extensor muscle strength and pain score of patients in conventional treatment+joint mobilization surgery+MET group were respectively significantly larger and lower than those in conventional treatment alone group and conventional treatment+joint mobilization surgery group (P values all <0.05). The elbow extensor muscle strength and pain score of patients in conventional treatment+joint mobilization surgery group were similar to those in conventional treatment alone group (P>0.05). The elbow joint range of motion and elbow extensor muscle strength (with t values of 9.37, 25.54, 28.71, 6.70, 7.20, and 7.01, respectively, P<0.05), elbow joint function scores and pain scores (with Z values of 3.15, 3.63, 3.93, 3.30, 3.52, and 3.84, respectively, P<0.05) of patients in conventional treatment alone group, conventional treatment+joint mobilization surgery group, and conventional treatment+joint mobilization surgery+MET group after two months of treatment were significantly improved compared with those before treatment. Conclusions: The combination of MET and Maitland joint mobilization surgery can effectively improve elbow joint range of motion, elbow joint function, elbow extensor muscle strength, and pain of patients with deep elbow joint burns, therefore it is worthy of promotion.

PMID:37805753 | DOI:10.3760/cma.j.cn501225-20220608-00228

Zhonghua Shao Shang Za Zhi. 2023 May 20;39(5):434-442. doi: 10.3760/cma.j.cn501225-20230206-00035.

ABSTRACT

Objective: To investigate the effect and mechanism of glycine on rat cardiomyocytes pretreated with serum from burned rats (hereinafter referred to as burn serum). Methods: Experimental research methods were adopted. Thirty gender equally balanced Wistar rats aged 7 to 8 weeks were collected, 10 of which were used to prepare normal rat serum (hereinafter referred to as normal serum), and the other 20 were inflicted with full-thickness burn of 30% total body surface area to prepare burn serum. Primary cardiomyocytes were isolated and cultured from the apical tissue of 180 Wistar rats aged 1 to 3 days by either gender for follow-up experiments. Cells were divided into normal serum group and burn serum group treated with corresponding serum according to the random number table (the same grouping method below). Trypanosoma blue staining was performed at post treatment hour (PTH) 1, 3, 6, 9, and 12 to detect the cell survival rate. Cells were divided into burn serum alone group treated with burn serum for 6 h followed by routine culture of 30 min and 0.4 mmol/L glycine group, 0.8 mmol/L glycine group, 1.2 mmol/L glycine group, 1.6 mmol/L glycine group, and 2.0 mmol/L glycine group treated with burn serum for 6 h followed by culture of 30 min with corresponding final molarity of glycine, i.e., at post intervention hour (PIH) 6.5, the cell survival rate was detected as before. Cells were divided into normal serum group, burn serum alone group, 0.8 mmol/L glycine group, 1.2 mmol/L glycine group, and 1.6 mmol/L glycine group, with the same intervention of 6.5 h as before, respectively. The content of adenosine monophosphate (AMP) and adenosine triphosphate (ATP) was detected by high performance liquid chromatography, and the AMP/ATP ratio was calculated. The protein expressions of phosphorylated mammalian target of rapamycin complex 1 (p-mTORC1), phosphorylated p70 ribosomal protein S6 kinase (p-p70 S6K), phosphorylated eukaryotic translation initiation factor 4E-binding protein 1 (p-4E-BP1), and phosphorylated AMP-activated protein kinase (p-AMPK) were detected by Western blotting. Cells were divided into normal serum group, burn serum alone group, 0.8 mmol/L glycine group intervened as before and 0.8 mmol/L glycine+25 ng/mL rapamycin group treated with burn serum followed by culture with two reagents. The expressions of heat shock protein 70 (HSP70), metallothionein (MT), and tubulin were detected by immunofluorescence method after 30 min of corresponding culture at PTH 1, 3, and 6, i.e., at PIH 1.5, 3.5, and 6.5, and the microtubule morphology was observed at PIH 6.5. The sample number at each time point was 10. Data were statistically analyzed with analysis of variance for factorial design, one-way analysis of variance, least significant difference (LSD)-t test, LSD test, and Bonferroni correction. Results: At PTH 1, 3, 6, 9, and 12, the cell survival rates in burn serum group were significantly lower than those in normal serum group (with t values of 4.96, 16.83, 35.51, 34.33, and 27.88, P<0.05). In burn serum group, the cell survival rate at PTH 3, 6, 9, or 12 was significantly lower than that at PTH 1 (P<0.05), the cell survival rate at PTH 6, 9, or 12 was significantly lower than that at PTH 3 (P<0.05), and the cell survival rate at PTH 6 was similar to that at PTH 9 (P>0.05) but significantly higher than that at PTH 12 (P<0.05). Treatment of 6 h was selected as the follow-up intervention time of burn serum. At PIH 6.5, compared with that in burn serum alone group, the cell survival rate in each glycine group was significantly increased (P<0.05). The cell survival rate in 0.8 mmol/L glycine group was the highest, and 0.8, 1.2, and 1.6 mmol/L were selected as subsequent glycine intervention concentrations. At PIH 6.5, the AMP/ATP ratio of cells in burn serum alone group was significantly higher than that in normal serum group, 1.2 mmol/L glycine group, or 1.6 mmol/L glycine group (P values all <0.05), and the AMP/ATP ratio of cells in 1.6 mmol/L glycine group was significantly lower than that in 0.8 mmol/L glycine group (P<0.05). At PIH 6.5, the protein expressions of p-mTORC1, p-p70 S6K, and p-4E-BP1 of cells in normal serum group, burn serum alone group, 0.8 mmol/L glycine group, 1.2 mmol/L glycine group, and 1.6 mmol/L glycine group were 1.001±0.037, 0.368±0.020, 1.153±0.019, 1.128±0.062, 1.028±0.037, 0.96±0.07, 0.63±0.12, 1.17±0.13, 1.13±0.16, 1.11±0.11, and 0.98±0.06, 0.45±0.08, 1.13±0.05, 0.77±0.12, 0.51±0.13. Compared with those in burn serum alone group, the protein expressions of p-mTORC1, p-p70 S6K, and p-4E-BP1 of cells in normal serum group and each glycine group were significantly increased (P<0.05), while the protein expressions of p-AMPK were significantly decreased (P<0.05). Compared with those in 0.8 mmol/L glycine group, the protein expression of p-4E-BP1 of cells in 1.2 mmol/L glycine group and the protein expressions of p-mTORC1 and p-4E-BP1 of cells in 1.6 mmol/L glycine group were significantly decreased (P<0.05). Compared with those in 1.2 mmol/L glycine group, the protein expressions of p-mTORC1 and p-4E-BP1 of cells in 1.6 mmol/L glycine group were significantly decreased (P<0.05), while the protein expression of p-AMPK was significantly increased (P<0.05). Compared with those in normal serum group, the expression of tubulin of cells in burn serum alone group was significantly decreased at PIH 1.5, 3.5, and 6.5 (P<0.05), while the expression of HSP70 of cells at PIH 1.5 and 3.5 and the expression of MT at PIH 3.5 and 6.5 were significantly increased (P<0.05). The expressions of HSP70 and MT of cells at PIH 1.5, 3.5, and 6.5 and the expression of tubulin at PIH 1.5 and 3.5 in burn serum alone group and 0.8 mmol/L glycine+25 ng/mL rapamycin group were significantly lower than those in 0.8 mmol/L glycine group (P<0.05). At PIH 6.5, compared with that in normal serum group, the cell microtubule structure in burn serum alone group was disordered; the cell boundary in 0.8 mmol/L glycine group was clearer than that in burn serum alone group, and the microtubule structure arranged neatly near the nucleus. Compared with that in 0.8 mmol/L glycine group, 0.8 mmol/L glycine+25 ng/mL rapamycin group had unclear cell boundaries and disordered microtubule structure. Conclusions: Burn serum can cause cardiomyocytes damage in rats. Glycine can significantly up-regulate mammalian target of rapamycin/p70 ribosomal protein S6 kinase/eukaryotic translation initiation factor 4E-binding protein 1 signaling pathway through AMP-activated protein kinase, promote the synthesis of protective proteins HSP70, MT, and tubulin, stabilize the microtubule structure, and exert cardiomyocytes protection function.

PMID:37805752 | DOI:10.3760/cma.j.cn501225-20230206-00035

Mol Oral Microbiol. 2023 Oct 7. doi: 10.1111/omi.12434. Online ahead of print.

ABSTRACT

The multi-batch reanalysis approach of jointly reevaluating gene/genome sequences from different works has gained particular relevance in the literature in recent years. The large amount of 16S ribosomal ribonucleic acid (rRNA) gene sequence data stored in public repositories and information in taxonomic databases of the same gene far exceeds that related to complete genomes. This review is intended to guide researchers new to studying microbiota, particularly the oral microbiota, using 16S rRNA gene sequencing and those who want to expand and update their knowledge to optimise their decision-making and improve their research results. First, we describe the advantages and disadvantages of using the 16S rRNA gene as a phylogenetic marker and the latest findings on the impact of primer pair selection on diversity and taxonomic assignment outcomes in oral microbiome studies. Strategies for primer selection based on these results are introduced. Second, we identified the key factors to consider in selecting the sequencing technology and platform. The process and particularities of the main steps for processing 16S rRNA gene-derived data are described in detail to enable researchers to choose the most appropriate bioinformatics pipeline and analysis methods based on the available evidence. We then produce an overview of the different types of advanced analyses, both the most widely used in the literature and the most recent approaches. Several indices, metrics and software for studying microbial communities are included, highlighting their advantages and disadvantages. Considering the principles of clinical metagenomics, we conclude that future research should focus on rigorous analytical approaches, such as developing predictive models to identify microbiome-based biomarkers to classify health and disease states. Finally, we address the batch effect concept and the microbiome-specific methods for accounting for or correcting them.

PMID:37804481 | DOI:10.1111/omi.12434