Associated predictors of prolonged length of stay in patients surviving extensive burns: A large multicenter retrospective study.

BACKGROUND: Patients with extensive burns are critically ill and have long treatment periods. Length of stay (LOS) is a good measure for assessing treatment. This study sought to identify predictors of prolonged LOS in patients with extensive burns (≥50% TBSA). METHODS: This retrospective multicenter cohort study included adults aged ≥ 18 years who survived extensive burns in three burn centers in Eastern China between January 2016 and June 2022. Epidemiological, demographic and clinical outcomes data were extracted from electronic medical records and compared between patients with/without prolonged LOS, which was defined as LOS greater than the median. Logistic regression analysis was used to identify predictors of prolonged LOS. RESULTS: The study sample included 321 patients, of whom 156 (48.6%) had an LOS of 58 days (IQR 41.0-77.0). Univariate regression analysis showed that increased total burn area and increased full-thickness burn area; electrical, chemical and other burns; increased erythrocytes, leukocytes, platelets or serum creatinine within 24 h of admission; concomitant inhalation injury, pulmonary edema, sepsis, bloodstream infection, wound infection, pulmonary infection, urinary tract infection, or HB < 70 g/L during hospitalization were associated with prolonged LOS in patients with extensive burns. Increased number of surgical operations, mechanical ventilation and renal replacement therapy were also associated with prolonged LOS (P < 0.05 or P < 0.001). Multivariate regression analysis revealed that increased total burn area (ratio 1.032, 95%CI 1.01-1.055; P = 0.004), electrical and chemical or other burns (3.282, 1.335-8.073; P = 0.01), development of wound infection (2.653 1.285-5.481; P = 0.008) and increased number of operative procedures (1.714, 1.388-2.116, P < 0.001) were significant predictors. CONCLUSIONS: Increased area of full-thickness burn,occurrence of electrical and chemical or other burns,occurrence of wound infection and increased number of surgeries are the best predictors of prolonged LOS in patients with extensive burns. Clarifying relevant predictors of burn patients' LOS provides a reliable reference for clinical treatment.

Differential miRNA expression profiles in human keratinocytes in response to protein kinase C inhibitor.

Aberrant expression of microRNAs (miRNAs) is widely accepted to be involved in keratinocyte differentiation and to be dependent on activation of the protein kinase C (PKC) pathway. However, the miRNA profiles and biological characteristics of keratinocytes induced by specific inhibitors of PKC have yet to be elucidated. The present study aimed to explore the differential miRNA expression profiles in keratinocytes treated with the PKC inhibitor GF109203X, by conducting a bioinformatics analysis. Parts of the GF109203X­induced keratinocytes formed distinct clones after 2 days of culture, and the expression of intergrin ß1, cytokeratin (CK)19 and CK14 were positive, whereas CK10 expression was negative. A total of 79 miRNAs were differentially expressed in keratinocytes treated with GF109203X, among which 45 miRNAs were upregulated and 34 were downregulated. The significantly upregulated microRNAs includedhsa­miR­1­3p and miR­181c­5p, whereas hsa­miR­31­5p and hsa­let­7c­3p were significantly downregulated. In addition, the results of reverse transcription­quantitative polymerase chain reaction exhibited consistency with the microarray results. An enrichment analysis demonstrated that certain target genes of the differentially expressed miRNAs serve an important role in cell proliferation and differentiation, cell cycle progression and apoptosis, etc. These results revealed that GF109203X induced the differential expression of certain miRNAs when keratinocytes began showing the characteristics of epidermal­like stem cells, which may provide a novel approach for wound healing and regeneration of skin tissues.

[Effects of recombinant human granulocyte-macrophage colony-stimulating factor on wound healing and microRNA expression in diabetic rats].

OBJECTIVE: To investigate the effects of recombinant human granulocyte macrophage colony-stimulating factor (rhGM-CSF) on wound healing and microRNA expression in diabetic rats. METHODS: Eighteen male SD rats of clean grade were used to reproduce diabetes model. Four weeks later, a total of 64 full-thickness skin wounds were created on the back of 16 rats with established diabetes, with 4 wounds on each rat. Two symmetrical wounds on either side of the spine were created as a pair according to paired design. Then the wounds were divided into groups A and B according to the random number table and blind method (red and blue tags on the rhGM-CSF or the gel vehicle), with 32 wounds in each group. The ointment with red tag was applied on the wounds of group A and the blue one on group B. The application was conducted once a day, with a thickness of 3 mm, up to post injury day (PID) 14. Gross observation of wound healing was conducted on PID 3, 7, 14. The wound healing rate was determined on PID 3 and 7. On PID 3, 7, 14, tissues from 2, 4, and 8 wounds were harvested from each group respectively for the observation of the histopathological changes with HE staining, and also for analyzing the expression of proliferating cell nuclear antigen (PCNA) and CD31 with immunohistochemical staining (denoted as absorbance value). On PID 7, tissues from 6 wounds in each group were harvested for microarray gene chip to screen the differentially expressed microRNAs. Enrichment analysis of Kyoto encyclopedia of genes and genomes (KEGG) signaling pathway on the differentially expressed microRNAs were performed after the microRNA screening results were validated by real-time fluorescent quantitative RT-PCR. Data were processed with paired t test or two-sample t test. RESULTS: (1) On PID 3, the wound area was significantly decreased, and the wound granulation was significantly proliferated in both groups. On PID 7, the wound area was further decreased, and the wound area was almost filled by granulation in both groups; the conditions in group A were better. On PID 14, all the wounds in group A were almost healed, while a small area of raw wound with incrustation still remained in some wounds of group B. On PID 3 and 7, the wound healing rates of group A were (41 ± 5)% and (75 ± 4)%, significantly higher than those of group B [(31 ± 9)% and (71 ± 4)%, with t values respectively 10.13 and 8.06, P values below 0.001]. (2) On PID 3, the epidermal cells, endothelial cells, and Fbs in the wounds of 2 groups were sparse, with heavy infiltration of inflammatory cells. The above condition in the wounds was better in group A than in group B. On PID 7, the epidermal cells, endothelial cells, and Fbs were gradually well arranged in group A; infiltration of inflammatory cells decreased, and the condition was better than that of group B. On PID 14, the wounds of group A were completely covered by epidermis, while infiltration of inflammatory cells still remained in some wounds of group B. (3) On PID 3, 7, 14, the positive expressions of CD31 and PCNA in group A were respectively 0.275 ± 0.018, 0.345 ± 0.034, 0.305 ± 0.023; 0.406 ± 0.063, 0.223 ± 0.011, 0.045 ± 0.022. They were significantly higher than those of group B (0.222 ± 0.020, 0.229 ± 0.018, 0.197 ± 0.015; 0.324 ± 0.039, 0.162 ± 0.012, 0.018 ± 0.020, with t values from 2.281 to 9.652, P < 0.05 or P < 0.01). (4) According to the microRNAs detection and screening, as compared with group B, 18 microRNAs were up-regulated while 13 were down-regulated in the wounds of group A. (5) The results of real-time fluorescent quantitative RT-PCR had good consistency with the results of microRNAs detection. (6) Enrichment analysis of KEGG signaling pathway showed that among the 31 differentially expressed microRNAs, 4 took part in the MAPK signaling pathway, 3 took part in the Wnt signaling pathway, 1 took part in the TGF-ß signaling pathway, 3 took part in the epidermal growth factor receptor signaling pathway, 2 took part in the cell cycle pathway, 5 took part in the axon guidance signaling pathway, 6 took part in the focal adhesion pathway, 3 took part in the regulation of actin cytoskeleton pathway, 1 took part in the extracellular cell matrix receptor pathway, 3 took part in the adherens junction pathway, and 1 took part in the cell adhesion molecules pathway. After disclosing the blind, it showed that the ointment with red tag was the rhGM-CSF gel and the blue one was gel vehicle. CONCLUSIONS: The rhGM-CSF gel can promote wound healing in diabetic rats, producing significant differential microRNA expression in wounds, and they may be the target at gene post-transcriptional level of rhGM-CSF gel in promoting wound healing.

[Effect of tetrandine on gene expression of collagen type I, collagen type III and TGF-beta1 in scar tissue's of rabbits ear].

OBJECTIVE: To observe the effect of tetrandine on gene expression of collagen type I, collagen type III, transformation growth factor-beta1 and to investigate the inhibitory effect of tetrandine on the scar tissue hyperplasia in rabbits' ears. METHODS: After the scar model was formed on the rabbits' ears, the rabbits were divided into 4 groups to receive intro-lesion injection with saline, or prednisolone (Pre) or tetrandrine in low concentration (L-Tet, 1.0 mg/ml) or tetrandrine in high concentration (H-Tet, 7.5 mg/ml). The morphological changes of scar tissue were observed. The changes of fibroblasts quantity and collagen expression were observed with HE and Masson staining. Immunohistochemical study was used to observe the expression level of collagen type I and collagen type III and TGF-beta1. Collagen type I and collagen type III and TGF-beta1, and signal factor Smad 3 mRNA were detected with RT-PCR. RESULTS: (1) 24 days after injury, all the wounds healed completely with formation of red, tough and hypertrophic scar. HE and Masson staining showed significant increase of fibroblasts and collagen density with irregularly arrangement. (2) Compared with that in saline group, the scar in other groups became softer, lighter and thinner, especially in H-Tet group. (3) HE and Masson staining shows the scar in Tet and Pre groups contained less fibroblasts and lower collagen dentsity with comparatively regular arrangement than that in saline group (P < 0.01), especially in H-Tet group. (4) According to the immunohistochemical study, the expression of collage type I and III and TGF-beta was positive in all the groups, but the positive rate and the ratio of collagen density I to III decreased in the order of saline, L-Tet, H-Tet and Pre groups (P < 0.01). (5) PT-PCR detection results showed that the amplification bands brightness of collagen type I and III and TGF-beta1 and signal molecular Smad 3 mRNA in scar tissue were obviously different. Compared with that in saline group, the expression of collagen type I and III and TGF-beta1 and Smad 3 mRNA decreased in Tet and Pre groups (P < 0.01). H-Tet group showed the most obvious reduce in the expression of type I collagen and TGF-beta1 and Smad 3 mRNA. Conclusions Tetrandine can significantly suppress the expression of collagen type I and collagen type III and TGF-beta1 on hypertrophic scar of rabbit ears, and reduce signal factor Smad 3 mRNA' s expression. It may be one of the important mechanism for its inhibitory effect on scar hyperplasia.

[Experimental study on the differentiation of human induced pluripotent stem cells into epidermal-like stem cells].

OBJECTIVE: To investigate the feasibility of differentiation of human induced pluripotent stem cells (iPSCs) into epidermal-like stem cells. METHODS: (1) Human strain of iPSCs were plated on-to trophoblast of inactivated Fb strain of mouse embryos and cultured in complete medium of embryonic stem cells, iPSCs were subcultured by collagenase IV digestion method. The morphology and growth of iPSCs were observed under inverted phase contrast microscope, and the cells were stained with alkaline phosphatase (AKP). iPSCs were cultured in incomplete medium of embryonic stem cells to observe the ability of embryoid body formation. (2) Human iPSCs were inoculated onto 6-well plate covered with human amniotic membrane to culture as induction group. Other iPSCs were cultured on 6-well plate without human amniotic membrane as control group. Morphological changes in iPSCs in two groups were observed. Expressions of integrin beta1 and CK19 of iPSCs in two groups were determined by immunocytochemical staining. RESULTS: Human iPSCs showed a typical stem cell clone-like growth with a clear boundary, and they proliferated vigorously in complete medium of embryonic stem cells. These cells were AKP-positive. iPSCs formed embryoid body in trophoblast-free and suspension culture conditions. After 4 days of co-culture, stem cell clones were formed on the surface of amniotic membrane in induction group, and part of the cells were integrin beta1 and CK19 positive. Most of the cells died, and no integrin beta1 and CK19 positive cells were found in control group. CONCLUSIONS: Human iPSCs can be differentiated into epidermal-like stem cells by amniotic membrane induction, and it lays an experimental basis for providing new source of seed cells of skin tissue engineering.

[Differential expression profile of microRNA between hyperplastic scar and normal skin].

OBJECTIVE: To explore the miRNA differential expression profiles of hyperplastic scar and normal skin so as to further elucidate the pathogenesis of hyperplastic scar and search for new therapeutic targets. METHODS: The total RNA was extracted from 5 human hyperplastic scar and normal skin tissues by Trizol. The specimens were collected from the First Affiliated Hospital of Nanchang University from November 2010 to May 2011, and purified by mirVana(TM) miRNA Isolation Kit and then labeled and hybridized by miRNA Complete Labeling and Hyb Kit. The images of hybridization were analyzed by the Feature Extraction (v10.7) software and the microarray results confirmed by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: In hyperplastic scar, 92 miRNA genes were up-regulated and 13 down-regulated. The most significantly up-regulated miRNAs were hsa-miR-564 and hsa-miR-936, etc. while hsa-miR-451, hsa-miR-223, hsa-miR-363 and hsa-miR-29b-1* became significantly down-regulated. The findings of RT-PCR on hsa-miR-21 and hsa-miR-451 of regulation were in a high concordance with the microarray results. CONCLUSION: Distinct differences of miRNA expression between human hyperplastic scar and normal skin, it may be closely correlated with the formation, development and evolution of hyperplastic scar.

[Screening of differential expression genes of human skin epidermal stem cells at different development stages by cDNA microarray technique].

OBJECTIVE: To analyze expression characteristics of human skin epidermal stem cell at different developmental stages, and to explore its biological significance. METHODS: Health skin samples from 28-32 w fetuses (F group), 4-12 y children (C group), and 35-55 y adult (A group) were harvested, with 10 cases in each group. Epidermis were separated using trypsin digestion and EDTA, and human epidermal stem cells were isolated and purified with type IV collagen attachment method. The monoclonal antibody of integrin beta1 and keratin 19 were used for detection and identification of epidermal stem cells by immunohistochemical staining. Total RNA was extracted from above cells by Trizol one-step method, and were detected by formaldehyde denaturing agarose gel electrophoresis. Probes were prepared and hybridized into cDNA microarray for scanning fluorescent signals and analysis of images, with two-fold differential expression value for screening. Significantly up/down-regulated genes were selected for verification by real time RT-PCR. RESULTS: By comparing expression profile between A and C groups, a total of 1808 genes with differential expression were detected, including 1089 up-regulated genes and 719 down-regulated genes, and they were classified into 128 categories. Among them, 1462 genes were known (found in GeneBank), 346 genes were unknown. A total of 4534 genes with differential expression were detected between C and F groups, in which 1783 genes were up-regulated and 2751 genes were down-regulated, and they were classified into 216 categories. Among them, 3577 genes were known (found in GeneBank), and 957 genes were unknown. There were 1104 genes with differential expression consistently detected in F, C and A groups, which were classified into 32 categories according to gene function. Among them, 94 genes were consistently up-regulated and 75 genes consistently down-regulated. Test results of real time RT-PCR were in accordance with above-mentioned results. CONCLUSIONS: Gene expression profiles of epidermal stem cells cultured in vitro, harvested from fetuses, children, and adult, exhibit obvious difference. This may be closely related to different stages of proliferation and differentiation of human epidermal stem cell and self-repair ability of wound at different developmental stages.