[Study on the consistency between CBCT image features of sphenopalatine foramen and those seen in nasal endoscopy].

Objective:To study the clinical anatomy of the sphenopalatine foramina by dissecting the sphenopalatine foramina during Vidian nerve branch neurotomy. The anatomy and CBCT images of sphenopalatine foramen were analyzed to facilitate the navigational of clinical operation using CBCT images. Methods:From October 2017 to September 2023, 84 cases（168 sides） of Vidian nerve branch neurotomy in our department were collected. The clinical summary was made according to the anatomy of sphenopalatine foramen during the operation. Preoperative CBCT imaging findings of the sphenopalatine foramina were also studied. Results:The clinical anatomy of sphenopalatine foramen could be divided into four types: middle meatus type（1.19%）, trans-meatus type（62.29%）, superior meatus type（33.33%） and double foramen type（1.19%）. The incidence of ethmoidal ridge was 98.81%. The distance from sphenopalatine foramina to posterior nasal canal were（14.63±2.66） mm to left and（14.65±2.63） mm to right, The position Angle ∠a of lower margin of sphenopalatine foramina were（62.36±10.05）° to left and（61.51±11.82）° to right, respectively. Axial CT images can be used to divide the sphenopalatine foramen into five levels: the upper edge of the sphenopalatine foramen level, the Vidian nerve level, the basal plate interaction level, the lower edge of the sphenopalatine foramen level and the pterygopalatine canal level. The agreement between endoscopic anatomy of sphenopalatine foramen and imaging navigation was 100%. Conclusion:The sphenopalatine foramina exhibit various anatomical types. The preoperative navigational CBCT reading can effectively identify the type of sphenopalatine foramina, guide the choice of surgical method, and help avoid serious complications. This has significant clinical application value.

PERP May Affect the Prognosis of Lung Adenocarcinoma by Inhibiting Apoptosis.

Background: PERP, a member of the peripheral myelin protein gene family, is a new therapeutic target in cancer. The relationships between PERP and immune cell infiltration in lung cancer have not been studied. Therefore, the role of PERP in the tumour microenvironment (TME) of lung cancer needs to be further explored. Methods: In this study, we explored the association between PERP expression and clinical characteristics by analysing data from the TCGA database. Cox regression and Kaplan‒Meier methods were used to investigate the relationship between the expression of PERP and overall survival in patients with lung adenocarcinoma (LUAD). The relationship between PERP expression and the degree of infiltration of specific immune cell subsets in LUAD was evaluated using the TIMER database and GEPIA. We also performed GO enrichment analysis and KEGG enrichment analysis to reveal genes coexpressed with PERP using the Coexpedia database. Finally, we verified the expression and function of PERP in LUAD tissues and the A549 cell line by RT‒PCR, Western blot, CCK-8, IHC, and wound healing assays. The mouse model was used to study the in vivo effects of PERP. Results: According to our results, PERP expression was significantly higher in LUAD tissues and associated with the clinical characteristics of the disease. Survival was independently associated with PERP in LUAD patients. We further verified that PERP might regulate B-cell infiltration in LUAD to affect the prognosis of LUAD. To identify PERP-related signalling pathways in LUAD, we performed a genome-aggregation analysis (GSEA) between low and high PERP expression datasets. LUAD cells express higher levels of PERP than paracarcinoma cells, and PERP inhibits the proliferation and metastasis of A549 cells through apoptosis. Conclusion: PERP may affect the prognosis of lung adenocarcinoma by inhibiting apoptosis and is associated with immune cell infiltration.

Comprehensive analysis of Eleutherococcus senticosus (Rupr. & Maxim.) Maxim. fruits based on UPLC-MS/MS and GC-MS: A rapid qualitative analysis.

Eleutherococcus senticosus (Rupr. & Maxim.) Maxim. fruits (ESF), as a natural edible fruit, has long been popularized. However, few studies have conducted comprehensive chemical analyses of it. This study aimed to assess nonvolatile, volatile, and fatty oil components of ESF and to preliminarily explore the antioxidant activities. The qualitative and quantitative analyses of volatile and fatty oil components of ESF from 15 different regions were performed by the gas chromatography-mass spectrometry (GC-MS). Totally, 37 and 28 compounds were identified from volatile oil and fatty oil, respectively. The ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS/MS) was used to accurately detect 43 compounds of nonvolatile components. The volatile and fatty oil components and nonvolatile components of ESF were used as samples to determine the antioxidant activity of 2,2-diphenyl-1-picrylhydrazyl (DPPH) in vitro. The components of ESF had antioxidant activity, and the nonvolatile components had stronger antioxidant activity. The results revealed that the proposed method, which is of great significance for the screening of new active ingredients, is valuable for the identification of pharmaceutical component and further development of food industry.

Integrated Nursing and Psychological Intervention for Tuberculosis Complicated by Lung Cancer: Clinical Efficacy.

Context: The incidence of tuberculosis (TB) complicated by lung cancer has been increasing yearly worldwide. The overlapping effects of these two diseases leads to difficulties in clinical treatment and care. Single-care modalities fail to meet the clinical-care requirements of these complex diseases for both psychological and physical treatment. Objective: The study intended to evaluate the clinical efficacy of integrated nursing plus a psychological intervention for patients with TB complicated by lung cancer. Design: The research team conducted a randomized controlled study. Setting: The study took place at the Affiliated Hospital of Hebei University in Baoding, Hebei, China. Participants: Participants were 60 patients with pulmonary TB complicated by lung cancer who received treatment at the hospital between January 2022 and December 2022. Interventions: The research team randomly assigned participants to one of two groups, each with 30 participants: (1) the control group, who received integrated nursing and (2) the intervention group who received integrated nursing plus a psychological intervention. Outcome Measures: The research team evaluated: (1) short-term clinical efficacy; (2) quality of life, using the Medical Outcomes Study's (MOS') 36-item Short-form Health Survey (SF-36); (3) levels of anxiety and depression, using the Self-Rating Anxiety Scale (SAS) and Self-Rating Depression Scale (SDS), respectively; and (4) nursing satisfaction. Results: No significant differences existed between the groups in demographic or clinical characteristics at baseline (P > .05). Compared to the control group, the intervention group's; (1) short-term clinical efficacy was significantly higher (P = .035); (2) scores on the SF-36 were significantly higher (all P < .001; (3) scores on the SAS and SDS were significantly lower (both P < .001); and (4) nursing satisfaction was significantly higher (P = .000). Conclusions: Integrated nursing plus psychological intervention can improve the quality of life of patients with TB complicated by lung cancer, alleviate their negative emotions, and enhance nursing satisfaction, thereby promoting patients' recoveries.

Rapid and Efficient Differentiation of Rodent Neural Stem Cells into Oligodendrocyte Progenitor Cells.

Oligodendrocyte progenitor cells (OPCs) may have beneficial effects in cell replacement therapy of neurodegenerative disease owing to their unique capability to differentiate into myelinogenic oligodendrocytes (OLs) in response to extrinsic signals. Therefore, it is of significance to establish an effective differentiation methodology to generate highly pure OPCs and OLs from some easily accessible stem cell sources. To achieve this goal, in this study, we present a rapid and efficient protocol for oligodendroglial lineage differentiation from mouse neural stem cells (NSCs), rat NSCs, or mouse embryonic stem cell-derived neuroepithelial stem cells. In a defined culture medium containing Smoothened Agonist, basic fibroblast growth factor, and platelet-derived growth factor-AA, OPCs could be generated from the above stem cells over a time course of 4-6 days, achieving a cell purity as high as ∼90%. In particular, these derived OPCs showed high expandability and could further differentiate into myelin basic protein-positive OLs within 3 days or alternatively into glial fibrillary acidic protein-positive astrocytes within 7 days. Furthermore, transplantation of rodent NSC-derived OPCs into injured spinal cord indicated that it is a feasible strategy to treat spinal cord injury. Our results suggest a differentiation strategy for robust production of OPCs and OLs from rodent stem cells, which could provide an abundant OPC source for spinal cord injury.

The expression of natriuretic peptide receptors in developing zebrafish embryos.

There are three isoforms of natriuretic peptide (NP) specific cell surface receptor: NP receptor-A (NPRA), receptor-B (NPRB), and receptor-C (NPRC). They are also known as NPR1, NPR2 and NPR3, respectively. NPs and their receptors were revealed to involve in diverse cellular and physiological processes including renal, cardiovascular, neuronal, and immunological aspects. However, the systematic analysis of the expression of these receptors in non-mammalian vertebrates is thus far lacking. In this study, two versions of the npr1 gene (npr1a and npr1b) in zebrafish was identified. Multiple sequences alignment analysis showed that zebrafish NPRs shared high homologies with NPRs of other species and possessed a typical signature domain of NPRs. The results of whole mount in situ hybridization and reverse transcription polymerase chain reaction analysis revealed that at embryonic stages, npr1a was mainly expressed in tectal ventricle, brian, heart and retina, whereas npr1b was broadly present in anterior pronephric duct. Unlike npr1, npr2 mainly expressed in branchial arches and neural tube during embryonic development. However, npr3 was expressed in pronephric ducts and corpuscle of stannius in zebrafish embryos at 72 hpf. In adults, we demonstrated that all the three NP receptors were highly existed in brain and kidney. Overall, these findings will provide an important basis for the functional analysis of NPs and its receptor during embryonic development.

MicroRNA-98 inhibits the cell proliferation of human hypertrophic scar fibroblasts via targeting Col1A1.

BACKGROUND: Hypertrophic scarring (HS) is a severe disease, and results from unusual wound healing. Col1A1 could promote the hypertrophic scar formation, and the expression of Col1A1 in HS tissue was markedly higher than that in the normal. In present study, we aimed to identify miRNAs as post-transcriptional regulators of Col1A1 in HS. METHODS: MicroRNA-98 was selected as the key miRNA comprised in HS. The mRNA levels of miR-98 in HS tissues and the matched normal skin tissues were determined by qRT-PCR. MTT and flow cytometry were used to determine the influence of miR-98 on cell proliferation and apoptosis of HSFBs, respectively. Col1A1 was found to be the target gene of miR-98 using luciferase reporter assay. Luciferase assay was performed to determine the relative luciferase activity in mimic NC, miR-98 mimic, inhibitor NC and miR-98 inhibitor with Col1A13'-UTR wt or Col1A13'-UTR mt reporter plasmids. The protein expression of Col1A1 in HSFBs after transfection with mimic NC, miR-98 mimic, inhibitor NC and miR-98 inhibitor were determined by western blotting. RESULTS: The mRNA level of miR-98 in HS tissues was much higher than that in the control. Transfection of HSFBs with a miR-98 mimic reduced the cell viability of HSFBs and increased the apoptosis portion of HSFBs, while inhibition of miR-98 increased cell viability and decreased apoptosis portion of HSFBs. miR-98 inhibitor increased the relative luciferase activity significantly when cotransfected with the Col1A1-UTR reporter plasmid, while the mutant reporter plasmid abolished the miR-98 inhibitor-mediated increase in luciferase activity. Western blotting revealed that overexpression of miR-98 decreased the expression of Col1A1. CONCLUSIONS: Overexpression of miR-98 repressed the proliferation of HSFBs by targeting Col1A1.

MicroRNA expression signature and the therapeutic effect of the microRNA­21 antagomir in hypertrophic scarring.

Hypertrophic scars (HS) area fibroproliferative disorder of the skin, which causes aesthetic and functional impairment. However, the molecular pathogenesis of this disease remains largely unknown and currently no efficient treatment exists. MicroRNAs (miRNAs) are involved in a variety of pathophysiological processes, however the role of miRNAs in HS development remains unclear. To investigate the miRNA expression signature of HS, microarray analysis was performed and 152 miRNAs were observed to be differentially expressed in HS tissue compared with normal skin tissues. Of the miRNAs identified, miRNA­21 (miR­21) was significantly increased in HS tissues and hypertrophic scar fibroblasts (HSFBs) as determined by reverse transcription­quantitative polymerase chain reaction analysis. It was also observed that, when miR­21 in HSFBs was blocked through use of an antagomir, the phenotype of fibrotic fibroblasts in vitro was reversed, as demonstrated by growth inhibition, induction of apoptosis and suppressed expression of fibrosis­associated genes collagen type I α 1 chain (COL1A1), COL1A2 and fibronectin. Furthermore, miR­21 antagomir administration significantly reduced the severity of HS formation and decreased collagen deposition in a rabbit ear HS model. The total scar area and scar elevation index were calculated and were demonstrated to be significantly decreased in the treatment group compared with control rabbits. These results indicated that the miR­21 antagomir has a therapeutic effect on HS and suggests that targeting miRNAs may be a successful and novel therapeutic strategy in the treatment of fibrotic diseases that are difficult to treat with existing methods.

A Selenium-Enriched Ziyang Green Tea Polysaccharide Induces Bax-Dependent Mitochondrial Apoptosis and Inhibits TGF-ß1-Stimulated Collagen Expression in Human Keloid Fibroblasts via NG2 Inactivation.

Keloids are fibroproliferative disorders characterized by the overabundant deposition of extracellular matrix (ECM), especially collagen and overgrowth of scar tissue in response to cutaneous injury. In this study, we isolated a selenium (Se)-containing polysaccharide (Se-ZGTP-I) from Ziyang green tea and explored its potential therapeutic effects on keloid fibroblasts formation. 3-(4,5-Dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and annexin V/propidium iodide (PI) staining assays demonstrated that Se-ZGTP-I or neuron-glia 2 (NG2) short hairpin RNA (shRNA) significantly inhibited proliferation of human keloid fibroblasts via induction of apoptosis. Besides, the activation of caspase-3 and the subsequent cleavage of poly (ADP-ribose) polymerase (PARP) were observed in keloid fibroblasts following Se-ZGTP-I (200 and 400 µg/ml) or NG2 shRNA treatment. Moreover, Western blotting analysis showed that treatment of keloid fibroblasts with Se-ZGTP-I (200 and 400 µg/ml) or NG2 shRNA resulted in an increase of pro-apoptotic protein Bax expression and a decrease in expression levels of anti-apoptotic protein Bcl-2 and NG2. In addition, type I collagen biosynthesis and protein expression in keloid fibroblasts following TGF-ß1 stimulation were decreased by Se-ZGTP-I (200 and 400 µg/ml) or NG2 shRNA management. Current findings imply that Se-ZGTP-I has a therapeutic potential to intervene and prevent keloid formation and other fibrotic diseases.

MicroRNA-98 inhibits the cell proliferation of human hypertrophic scar fibroblasts via targeting Col1A1

BACKGROUND: Hypertrophic scarring (HS) is a severe disease, and results from unusual wound healing. Col1A1 could promote the hypertrophic scar formation, and the expression of Col1A1 in HS tissue was markedly higher than that in the normal. In present study, we aimed to identify miRNAs as post-transcriptional regulators of Col1A1 in HS. METHODS: MicroRNA-98 was selected as the key miRNA comprised in HS. The mRNA levels of miR-98 in HS tissues and the matched normal skin tissues were determined by qRT-PCR. MTT and flow cytometry were used to determine the influence of miR-98 on cell proliferation and apoptosis of HSFBs, respectively. Col1A1 was found to be the target gene of miR-98 using luciferase reporter assay. Luciferase assay was performed to determine the relative luciferase activity in mimic NC, miR-98 mimic, inhibitor NC and miR-98 inhibitor with Col1A13'-UTR wt or Col1A13'-UTR mt reporter plasmids. The protein expression of Col1A1 in HSFBs after transfection with mimic NC, miR-98 mimic, inhibitor NC and miR-98 inhibitor were determined by western blotting. RESULTS: The mRNA level of miR-98 in HS tissues was much lower than that in the control. Transfection of HSFBs with a miR-98 mimic reduced the cell viability of HSFBs and increased the apoptosis portion of HSFBs, while inhibition of miR-98 increased cell viability and decreased apoptosis portion of HSFBs. miR-98 inhibitor increased the relative luciferase activity significantly when cotransfected with the Col1A1-UTR reporter plasmid, while the mutant reporter plasmid abolished the miR-98 inhibitor-mediated increase in luciferase activity. Western blotting revealed that overex-pression of miR-98 decreased the expression of Col1A1. CONCLUSIONS: Overexpression of miR-98 repressed the proliferation of HSFBs by targeting Col1A1.

Combretastatin A-4 efficiently inhibits angiogenesis and induces neuronal apoptosis in zebrafish.

Cis-stilbene combretastatin A-4 (CA-4) and a large group of its derivant compounds have been shown significant anti-angiogenesis activity. However the side effects even the toxicities of these chemicals were not evaluated adequately. The zebrafish model has become an important vertebrate model for evaluating drug effects. The testing of CA-4 on zebrafish is so far lacking and assessment of CA-4 on this model will provide with new insights of understanding the function of CA-4 on angiogenesis, the toxicities and side effects of CA-4. We discovered that 7-9 ng/ml CA-4 treatments resulted in developmental retardation and morphological malformation, and led to potent angiogenic defects in zebrafish embryos. Next, we demonstrated that intraperitoneal injection of 5, 10 and 20 mg/kg CA-4 obviously inhibited vessel plexus formation in regenerated pectoral fins of adult zebrafish. Interestingly, we proved that CA-4 treatment induced significant cell apoptosis in central nervous system of zebrafish embryos and adults. Furthermore, it was demonstrated that the neuronal apoptosis induced by CA-4 treatment was alleviated in p53 mutants. In addition, notch1a was up-regulated in CA-4 treated embryos, and inhibition of Notch signaling by DAPT partially rescued the apoptosis in zebrafish central nervous system caused by CA-4.

Expression Profile of Long Noncoding RNAs in Human Earlobe Keloids: A Microarray Analysis.

Background. Long noncoding RNAs (lncRNAs) play key roles in a wide range of biological processes and their deregulation results in human disease, including keloids. Earlobe keloid is a type of pathological skin scar, and the molecular pathogenesis of this disease remains largely unknown. Methods. In this study, microarray analysis was used to determine the expression profiles of lncRNAs and mRNAs between 3 pairs of earlobe keloid and normal specimens. Gene Ontology (GO) categories and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed to identify the main functions of the differentially expressed genes and earlobe keloid-related pathways. Results. A total of 2068 lncRNAs and 1511 mRNAs were differentially expressed between earlobe keloid and normal tissues. Among them, 1290 lncRNAs and 1092 mRNAs were upregulated, and 778 lncRNAs and 419 mRNAs were downregulated. Pathway analysis revealed that 24 pathways were correlated to the upregulated transcripts, while 11 pathways were associated with the downregulated transcripts. Conclusion. We characterized the expression profiles of lncRNA and mRNA in earlobe keloids and suggest that lncRNAs may serve as diagnostic biomarkers for the therapy of earlobe keloid.

High glucose inhibits ClC-2 chloride channels and attenuates cell migration of rat keratinocytes.

BACKGROUND: Accumulating evidence has demonstrated that migration of keratinocytes is critical to wound epithelialization, and defects of this function result in chronic delayed-healing wounds in diabetes mellitus patients, and the migration has been proved to be associated with volume-activated chloride channels. The aim of the study is to investigate the effects of high glucose (HG, 25 mM) on ClC-2 chloride channels and cell migration of keratinocytes. METHODS: Newborn Sprague Dawley rats were used to isolate and culture the keratinocyte in this study. Immunofluorescence assay, real-time polymerase chain reaction, and Western blot assay were used to examine the expression of ClC-2 protein or mRNA. Scratch wound assay was used to measure the migratory ability of keratinocytes. Transwell cell migration assay was used to measure the invasion and migration of keratinocytes. Recombinant lentivirus vectors were established and transducted to keratinocytes. Whole-cell patch clamp was used to perform the electrophysiological studies. RESULTS: We found that the expression of ClC-2 was significantly inhibited when keratinocytes were exposed to a HG (25 mM) medium, accompanied by the decline of volume-activated Cl(-) current (I Cl,vol), migration potential, and phosphorylated PI3K as compared to control group. When knockdown of ClC-2 by RNAi or pretreatment with wortmannin, similar results were observed, including I Cl,vol and migration keratinocytes were inhibited. CONCLUSION: Our study proved that HG inhibited ClC-2 chloride channels and attenuated cell migration of rat keratinocytes via inhibiting PI3K signaling.

Stromal cell-derived factor 1 (SDF-1) accelerated skin wound healing by promoting the migration and proliferation of epidermal stem cells.

Epidermal stem cells could contribute to skin repair through the migration of cells from the neighboring uninjured epidermis, infundibulum, hair follicle, or sebaceous gland. However, little is known about the factors responsible for the complex biological processes in wound healing. Herein, we will show that the attracting chemokine, SDF-1/CXCR4, is a major regulator involved in the migration of epidermal stem cells during wound repair. We found that the SDF-1 levels were markedly increased at the wound margins following injury and CXCR4 expressed in epidermal stem cells and proliferating epithelial cells. Blocking the SDF-1/CXCR4 axis resulted in a significant reduction in epidermal stem cell migration toward SDF-1 in vitro and delayed wound healing in vivo, while an SDF-1 treatment enhanced epidermal stem cell migration and proliferation and accelerated wound healing. These results provide direct evidence that SDF-1 promotes epidermal stem cell migration, accelerates skin regeneration, and makes the development of new regenerative therapeutic strategies for wound healing possible.

PTEN inhibits proliferation and functions of hypertrophic scar fibroblasts.

Hypertrophic scar (HS) remains a major problem in plastic surgery. In order to explore the regulative effect of phosphatase and tensin homolog (PTEN) on HS, PTEN and AKT expression was detected by reverse transcription PCR, immunohistochemistry and western blot. Adenovirus-mediated PTEN overexpression in cultured hypertrophic scar fibroblasts (HSFBs) and normal skin fibroblasts was also introduced to evaluate its biological function. Our results showed that PTEN expression was significantly decreased in HS whereas p-Akt level was significantly higher in HS compared with normal skin (P < 0.01). Furthermore, we found that adenovirus-mediated PTEN overexpression led to decreased AKT activation, and significantly reduced cell proliferation and collagen synthesis of HSFBs, while increased the apoptosis. Taken together, these data suggest that PTEN inhibits proliferation and function of HSFBs through AKT pathway. Our results reveal a novel biological role for PTEN/AKT pathway in HS and suggest PTEN as a potential therapeutic target for HS treatment.

[Modulatory effect of Rac1 protein on epidermal stem cells migration during wound healing].

OBJECTIVE: To investigate modulatory role of Rac1 protein in epidermal stem cell (ESC) migration during wound healing, in order to provide a reference for enriching basic theory of wound healing and guiding clinical application. METHODS: Constitutively active mutant of Rac1 protein (Rac1Q61L) or dominant negative isoform of Rac1 protein (Rac1T17N) was transfected into ESC using a retroviral vector FUGW, and retroviral vector FUGW transfected into ESC in singles was used as blank control. The cells were divided into 3 parts according to the random number table and treated as follows. First, equal numbers of cells were inoculated into 24-well plates coated with collagen I (20 µg/mL), collagen IV (20 µg/mL) or fibronectin (10 µg/mL). Cells adhered to above matrices were quantitated using CytoTox 96 colorimetric kit. Second, 1000 cells adhered to collagen IV, after being stained with tetramethyl rhodamine isothiocyanate-phalloidin, were collected for observation of cell morphology and comparison of spreading area under confocal laser scanning microscope. Third, ESC with density of 2 × 10(5) cells per well were placed in upper compartment of Transwell chamber, DK-SFM culture medium alone or that containing stromal cell derived factor 1 (SDF-1) was added into lower compartment of Transwell chamber. Migration of ESC was observed using inverted phase contrast microscope, and the result was denoted as migration rate. Lastly, ESC with density of 7.5 × 10(5) cells per well was inoculated into 6-well plates for 12 hours, and treated with 4 µg/mL mitomycin C for 2 hours. The remaining scratch width of monolayer was respectively measured 6 hours or 12 hours after scratching to calculate the percentage of remaining scratch width. Data were processed with t test. RESULTS: Compared with that of blank control, the number of Rac1Q61L-transfected cells adhered to collagen I was significantly increased (t = 5.302,P < 0.05), while the number of Rac1T17N-transfected cells adhered to collagen I, IV, and fibronectin were all obviously decreased (with t value respectively 13.741, 15.676, 8.256, P values all below 0.05). Confocal laser scanning microscope showed that spreading area of Rac1Q61L-transfected ESC (with laminate pseudopodia on edge) and Rac1T17N-transfected ESC was respectively larger and smaller as compared with that of blank control. With SDF-1 effect, the migration rate of Rac1T17N-transfected ESC was decreased by 78.0% and Rac1Q61L-transfected ESC was increased by 43.4% as compared with that of blank control. Without SDF-1 effect, the migration rate of Rac1T17N-transfected ESC was decreased by 55.2%, while the migration rate of Rac1Q61L-transfected ESC was close to that of blank control. Six or 12 hours after scratching, the percentage of remaining scratch width in Rac1Q61L-transfected ESC was lower as compared with that in blank control [(39 ± 9)% vs. (43 ± 5)%, (6 ± 5)% vs. (18 ± 7)%, with t value respectively 1.027, 4.389, with P value respectively above and below 0.05], while that in Rac1T17N-transfected ESC [(81 ± 9)%, (71 ± 11)%, respectively] was obviously higher as compared with that in blank control (with t value respectively 11.386, 11.726, P values all below 0.05). CONCLUSIONS: Rac1 protein may control the migration of ESC by regulating its adhesion, spreading, and chemotaxis, and it plays an active role in wound healing accelerated by ESC.

[Promotion effect of stromal cell-derived factor 1 on the migration of epidermal stem cells in the healing process of frostbite-wound model ex vivo].

OBJECTIVE: To study the promotion effect of stromal cell-derived factor 1 (SDF-1) on the migration of epidermal stem cells (ESC) in the healing process of frostbite-wound model ex vivo. METHODS: A three-dimensional model of full-thickness frostbite of skin was constructed (with slot-like wound) out of skin equivalent. The expression of SDF-1 in wound stroma was observed with immunohistochemistry staining on post injury days (PID) 3 and 7. The model frostbite wounds were divided into control group (treated with PBS 50 microL per wound), SDF-1 group (treated with 100 ng/mL SDF-1, 50 microL per wound), and AMD3100 group [treated with 100 ng/mL AMD3100 (50 microL per wound) for 30 minutes, and then SDF-1 50 microL was added per wound]. The redistribution of ESC around wound was observed. RESULTS: The expression of SDF-1 in wound stroma increased gradually on PID 3 and 7. Compared with those in control and AMD3100 groups, there were more ESC and epithelial cell layers, and more integrin beta(1)-positive cells appeared at the basal layer of wound in SDF-1 group, and some of the positive cells migrated upward to epidermis. CONCLUSIONS: SDF-1 contributes to wound repair through promoting ESC to migrate toward and gather around wound edge. This may be one of the mechanisms of ESC participating in wound repair.

Small Rho GTPase Rac1 determines human epidermal stem cell fate in vitro.

Epidermal stem cells (ESCs) are essential not only for tissue homeostasis but also for skin to respond to insults, but the mechanisms of stem cell regulation are unknown. To investigate the function of Rac1 in ESC development, we introduced either the dominant negative isoform or constitutively the active mutant of Rac1 in cultured human ESCs by using a retroviral vector and then analyzed the consequences. Upon activation, Rac1 increased surface alpha6/beta1 integrin levels and promoted colony forming efficiency of ESCs. Conversely, dominant negative Rac1 caused a progressive reduction in growth rate, an inhibition of adhesiveness and a marked stimulation of terminal differentiation, without any effect on the cell cycle. These results were consistent with the role of Rac1 in determining the fate of ESCs by controlling their exit from the stem cell compartment. Our results reveal a novel biological role for Rac1 and provide new insights into the mechanism regulating ESCs.

The GTPase Rab3b/3c-positive recycling vesicles are involved in cross-presentation in dendritic cells.

Antigen cross-presentation in dendritic cells is a complex intracellular membrane transport process, but the underlying molecular mechanisms remain to be thoroughly investigated. In this study, we examined the effect of siRNA-mediated knockdown of 57 Rab GTPases, the key regulators of membrane trafficking, on antigen cross-presentation. Twelve Rab GTPases were identified to be associated with antigen cross-presentation, and Rab3b/3c was indicated to be colocalized with MHC class I molecules at perinuclear tubular structure. Tracing with fluorescence protein-tagged beta(2)-microglobulin demonstrated that the MHC class I molecules were internalized from the plasma membrane to Rab3b/3c-positive compartments, which were also colocalized with the internalized transferrin. Moreover, depletion of Rab3b/3c strongly reduced the fast phase recycling rate of transferrin receptors. Furthermore, the Rab3b/3c-positive compartments were colocalized with a fraction of Rab27a at a juxtaposition of phagosomes. Together, these data demonstrate that Rab3b/3c-positive recycling vesicles are involved in and may constitute one of the recycling compartments in exogenous antigen cross-presentation.