[Some thoughts on the research of mesenchymal stem cell exosomes and wound microenvironment].

Since researchers have found that the conditioned medium and exosomes of mesenchymal stem cells (MSCs) had the biological effects equivalent to those of MSCs, MSC exosomes (MSC-Exos), the representative product of MSCs' paracrine effect, have become the research focus of the "cell-free" therapy of MSCs. However, most researchers currently use conventional culture condition to culture MSCs and then isolate exosomes for the treatment of wound or other diseases. Theoretically, the paracrine effect of MSCs is directly associated with the pathological condition of the wound (disease) microenvironment or in vitro culture condition, and their paracrine components and biological effects may be altered with the changes of the wound (disease) microenvironment or in vitro culture condition. Thus, the feasibility of using traditional culture condition to culture MSCs for exosome extraction for the treatment of different diseases without considering the actual situation of the disease to be treated needs further discussion. Therefore, the author suggests that the research of MSC-Exos should consider the microenvironment of the wound (disease) to be treated. as much as possible, otherwise the extracted MSC-Exos may not be "accurate" or may not really achieve the treatment effect of MSCs. In this article, we summarized some thoughts of the author and problems related to the researches about MSC-Exos and wound microenvironment, and hoped to discuss with researchers.

A study of microRNA-223 in evaluating platelet reactivity in patients with acute ischemic stroke.

To investigate the relationship between plasma microRNA-223 expression and platelet reactivity in patients with acute ischemic stroke (AIS) and to evaluate its predictive value in clopidogrel resistance or high on-treatment platelet reactivity (HTPR). A total of 120 patients with acute ischemic stroke were screened in this study, and 60 patients were included in the acute ischemic stroke group according to the inclusion criteria and platelet reactivity after clopidogrel treatment. control group was 60 non-ischemic stroke patients hospitalized. The levels of phosphorylation of vasodilator-stimulated phosphoprotein (VASP) and adenosine diphosphate-induced platelet aggregation (ADP-PAg) in platelets were detected by flow cytometry. The expression level of plasma microRNA-223 was detected before and after clopidogrel treatment using quantitative real-time polymerase chain reaction (PcR). The AIS group was then divided into the clopidogrel non-HTPR and the clopidogrel HTPR groups based on the relative inhibition rate. We found that: 1) the VASP platelet reactivity index (PRI) was positively correlated with ADP-PAg; 2) before administration, the plasma microRNA-223 expression level and VASP-PRI were higher in the AIS group than in the control group; 3) after administration, the expression level of microRNA-223 was negatively correlated with VASP-PRI; 4) before and after treatment, the plasma microRNA-223 expression level in the clopidogrel HTPR group was lower than in the non-AIS patients; 5) before treatment, there was an interaction between the expression level of microRNA-223 in the plasma and the cYP2c19 loss-of-function (LOF) allele. The study showed that decreased plasma microRNA-223 expression levels in AIS patients indicate an increased risk of clopidogrel HTPR. carrying cYP2c19 LOF alleles may result in the microRNA-223 expression being more distinct. The combined detection of plasma microRNA-223 and cYP2c19 gene polymorphisms may be effective in predicting the occurrence of clopidogrel HTPR in patients with AIS.

[Clinical application effects of two longitudes three transverses method in perforator location of thoracodorsal artery perforator flap and deep wound repair].

Objective: To explore the clinical application value of two longitudes three transverses method in the location of the perforator of thoracodorsal artery perforator and deep wound repair. Methods: The retrospectively observational study was conducted. From December 2018 to June 2020, 17 patients with deep wounds who were admitted to the Affiliated Hospital of Zunyi Medical University met the inclusion criteria and were included in this study, including 7 males and 10 females, aged 12 to 72 years. The wound areas of patients after debridement were 7 cm×3 cm to 11 cm×7 cm. Two longitudinal lines were located through the midpoint of the armpit, the posterior superior iliac spine, and the protruding point of the sacroiliac joint, and three transverse lines were located 5, 10, and 15 cm below the midpoint of the armpit between the two longitudinal lines, i.e. two longitudes three transverses method, resulting in two trapezoidal areas. And then the thoracodorsal artery perforators in two trapezoidal areas were explored by the portable Doppler blood flow detector. On this account, a single or lobulated free thoracodorsal artery perforator flap or flap that carrying partial latissimus dorsi muscle, with an area of 7 cm×4 cm to 12 cm×8 cm was designed and harvested to repair the wound. The donor sites were all closed by suturing directly. The number and location of thoracodorsal artery perforators, and the distance from the position where the first perforator (the perforator closest to the axillary apex) exits the muscle to the lateral border of the latissimus dorsi in preoperative localization and intraoperative exploration, the diameter of thoracodorsal artery perforator measured during operation, and the flap types were recorded. The survivals of flaps and appearances of donor sites were followed up. Results: The number and location of thoracodorsal artery perforators located before operation in each patient were consistent with the results of intraoperative exploration. A total of 42 perforators were found in two trapezoidal areas, with 2 or 3 perforators each patient. The perforators were all located in two trapezoid areas, and a stable perforator (the first perforator) was located and detected in the first trapezoidal area. There were averagely 1.47 perforators in the second trapezoidal area. The position where the first perforator exits the muscle was 2.1-3.1 cm away from the lateral border of the latissimus dorsi. The diameters of thoracodorsal artery perforators were 0.4-0.6 mm. In this group, 12 cases were repaired with single thoracodorsal artery perforator flap, 3 cases with lobulated thoracodorsal artery perforator flap, and 2 cases with thoracodorsal artery perforator flap carrying partial latissimus dorsi muscle. The patients were followed up for 6 to 16 months. All the 17 flaps survived with good elasticity, blood circulation, and soft texture. Only linear scar was left in the donor area. Conclusions: The two longitudes three transverses method is helpful to locate the perforator of thoracodorsal artery perforator flap. The method is simple and reliable. The thoracodorsal artery perforator flap designed and harvested based on this method has good clinical effects in repairing deep wound, with minimal donor site damage.

[Plasmids carried by carbapenems-resistant Klebsiella pneumoniae in burn patients and its correlation with strain transmission].

Objective: To explore the carrier status of carbapenems-resistant Klebsiella pneumoniae (CRKP) plasmids in burn patients and analyze the correlation of these plasmids with the transmission of CRKP. Methods: A retrospective observational study was conducted. A total of 26 CRKP strains, which were isolated from the clinic-related samples of 22 burn patients (with 20 males and 2 females, aged (42±16) years) admitted to the First Affiliated Hospital of Army Medical University (the Third Military Medical University) from January to December 2017, were collected and individually numbered. The plasmids of the strains were extracted by alkali lysis. After determination of the plasmid concentration by a nucleic acid concentration detector, the agarose gel electrophoresis was used to visualize the bands, and rough plasmids typing was performed. The plasmid of the smallest numbered CRKP in each plasmid type was transformed into competent Escherichia coli (E. coli) strain Top10 (hereinafter referred to as TOP10 strain). The growth of each transformed strains and a Top10 strain cultivated in ampicillin containing Luria-Bertani (LB) agar medium overnight was observed, and the proportion of successful transformation was calculated. The plasmids from the smallest numbered plasmid carrying CRKP strain of successfully transformed Top10 strains (hereinafter referred to as the smallest successfully transformed strain) and correspondingly numbered CRKP were extracted, and then, the agarose gel electrophoresis was used to visualize the bands. Aforementioned successfully transformed strains and a TOP10 strain were used for the antimicrobial susceptibility testing with 17 antibiotics commonly used in clinic. The plasmid from the smallest successfully transformed strain was sequenced using the next-generation sequencing technology. Bioinformatics analyses such as protein-coding gene prediction and protein sequence alignment were performed successively. The sequence was subsequently named pKP03-NDM1 according to the carrying of drug resistance gene. According to the whole genome sequence of the plasmid carried by the smallest successfully transformed strain, the polymerase chain reaction, agarose gel electrophoresis, and gene sequencing were used to detect the New Delhi metallo-beta lactamase-1 (blaNDM-1) of plasmids in the remaining 25 strains of CRKP. The ST typing in multilocus sequence typing of 26 strains of CRKP was analyzed based on the literature. Results: Plasmids were successfully extracted from 26 CRKP, with mass concentrations ranging from 19.3 to 189.8 ng/µL. Each of the 26 CRKP carrying plasmids showed at least one band longer than 2 500 bp in the agarose gel electrophoresis, which were roughly divided into 6 patterns of A, B, C, D, E, and F. After overnight cultivation, no growth of strains was observed in LB agar medium containing ampicillin inoculated with the TOP10 strain or TOP10 strains transformed by the plasmid of CRKP patterning A, B, D, or E. In contrast, TOP10 strains transformed by the pattern C plasmid from NO.3 CRKP and the pattern F plasmid from NO.15 CRKP resulted in numerous colony growths, and those transformed strains were named as TOP10-pKP03 and TOP10-pKP15, respectively. The proportion of successful transformation was 1/3. The plasmid carried by TOP10-pKP03 showed a single band in the agarose gel electrophoresis, which was the same size as the largest band of the plasmid from NO.3 CRKP. The TOP10 strain was sensitive to the 17 antibiotics commonly used in clinic. TOP10-pKP03 and TOP10-pKP15 were resistant to penicillins, cephalosporins, and carbapenems but remained sensitive to monocyclic ß-lactam, aminoglycosides, quinolones and tigecycline. The full length of the plasmid carried by TOP10-pKP03 was 41 190 bp. In addition to blaNDM-1, this plasmid carried bleMBL, T4SS, bleomycin resistance gene, conjugation transfer elements, and relaxase, etc. The plasmid showed 99% nucleotide identity similarity and the same length to the plasmid pJN24NDM1 extracted from an E. coli isolate JN24. Totally 16 (61.5%) CRKP were confirmed to carrying blaNDM-1 gene, among the ST typing of the 16 strains, 11 strains were ST11, while ST215, ST260, ST395, ST2230, and new ST had 1 strain each. Among the ST typing of 10 blaNDM-1-negative CRKP, 8 strains were ST11, while ST395 and ST2230 had 1 strain each. Conclusions: A blaNDM-1 gene carrying plasmid pKP03-NDM1 was extracted and sequenced from CRKP isolated from burn patients, with a high plasmid carrying rate. Meanwhile, this plasmid may mediate inter-CRKP and CRKP-E. coli horizontal transfer of blaNDM-1, leading to transmission of antimicrobial resistance.

[Research advances on the roles of exosomes derived from mesenchymal stem cells in wound healing and prevention and treatment of hypertrophic scars].

Skin is an important defense barrier of human body and one of the most vulnerable organs. Wounds are the result of damage to the integrity of skin. Chronic wounds and hypertrophic scar formation are the results of abnormal wound healing, and are also the clinical problems those need to be resolved urgently in the field of wound repair. In recent years, researchers have found that mesenchymal stem cells (MSCs) can promote wound healing, improve wound healing quality, and reduce scar formation. The therapeutic effect of MSCs may be derived from the exosomes derived from them. This paper reviews the research advances of exosomes derived from MSCs in wound healing and prevention and treatment of hypertrophic scars in recent years and looks up to the prospect for the clinical application.

Intrinsical localization of both topological (anti-kink) envelope and gray (black) gap solitons of the condensed bosons in deep optical lattices.

By developing quasi-discrete multiple-scale method combined with tight-binding approximation, a novel quadratic Riccati differential equation is first derived for the soliton dynamics of the condensed bosons trapped in the optical lattices. For a lack of exact solutions, the trial solutions of the Riccati equation have been analytically explored for the condensed bosons with various scattering length as. When the lattice depth is rather shallow, the results of sub-fundamental gap solitons are in qualitative agreement with the experimental observation. For the deeper lattice potentials, we predict that in the case of as>0, some novel intrinsically localized modes of symmetrical envelope, topological (kink) envelope, and anti-kink envelope solitons can be observed within the bandgap in the system, of which the amplitude increases with the increasing lattice spacing and (or) depth. In the case of as<0, the bandgap brings out intrinsically localized gray or black soliton. This well provides experimental protocols to realize transformation between the gray and black solitons by reducing light intensity of the laser beams forming optical lattice.

[Fetal cardiac intervention and perioperative management of fetus with hypoplastic right heart syndrome].

Objective: To summarize the experience of perioperative management strategy of fetal pulmonary valvuloplasty (FPV) for hypoplastic right heart syndrome (HRHS). Methods: In the retrospective study of perioperative data, 13 fetuses of HRHS were treated with FPV in Qingdao Women and Children's Hospital from July 2018 to June 2019. Results: (1) The evaluation indexes of the right ventricle in 13 fetuses before FPV: the mean ratio of tricuspid/mitral annulus, right/left ventricular length, pulmonary/aortic annulus, and tricuspid inflow time/cardiac cycle were 0.81±0.04, 0.56±0.14, 0.69±0.06, and 0.35±0.03, respectively. (2) All pregnant mothers underwent general anesthesia. The basic fetal heart rate was (156±12) beats per minutes before FPV, and 9 fetuses showed bradycardia during the operation, and recovered to normal after drug resuscitation. On the first day after FPV, two cases had bradycardia and pregnancy was terminated. The fluctuation of systolic blood pressure of pregnant mother was less than 20%, and there was no significant difference between preoperative and intraoperative pulse pressure [(36.0±5.6) vs (35.8±6.9) mmHg (1 mmHg=0.133 kPa); t=8.102, P=0.951]. (3) All cases of HRHS fetus successfully underwent FPV. The average gestational age was (27.3±0.8) weeks. The average operation time was (23.2±1.0) minutes. The ratio of tricuspid to mitral annulus (t=-2.513, P=0.022) and the ratio of right to left ventricular length (t=-3.373, P=0.003) were significantly improved at 6 weeks postoperatively. Ten fetuses were delivered, and there was no death after early intervention. (4) Of 13 pregnant women, 3 cases were nausea and vomiting on the day of FPV operation, the treatment of the symptoms was improved by tropisetron; one case had tolerable abdominal pain and improved without special treatment. Pregnant women had no major complications such as cardiac failure, abortion and death. (5) Chromosome karyotype analysis and microarray analysis of amniotic fluid was retained during the operation. No typical chromosome abnormality or other abnormal genetic diagnosis was found. Conclusions: FPV colud be used as an effective intervention measure to promote the development of right ventricle in HRHS fetuses. The scientific management of multidisciplinary professional technical team in perioperative period is particularly important to ensure the success of FPV and the safety of pregnant women and fetuses.

[Research on feasibility of in vitro inflammatory wound microenvironment simulated by using inflammatory wound tissue homogenate of mice].

Objective: To investigate the feasibility of in vitro inflammatory wound microenvironment simulated by using inflammatory wound tissue homogenate of mice. Methods: (1) Ten eight-week-old C57BL/6 male mice were collected and full-thickness skin tissue with diameter of 1.0 cm on both sides of the midline of the back was taken with a perforator to make the normal skin tissue homogenate supernatant. At 48 h after the full-thickness skin defect wound was established, the wound tissue within 2 mm from the wound edge was taken to make inflammatory wound tissue homogenate supernatant. Two kinds of tissue homogenate supernatant were taken to adjust the total protein concentration to 1 mg/mL, and the tumor necrosis factor α (TNF-α) content was detected by enzyme-linked immunosorbent assay. The number of sample was 6. (2) The primary passage of human umbilical cord mesenchymal stem cells (hUCMSCs) were collected and cultured to the 3rd passage with the normal exosomes being extracted from the hUCMSCs after cultured for 48 h. Another batch of hUCMSCs in the 3rd passage was collected and stimulated with inflammatory wound tissue homogenate supernatant of 30, 50, and 100 µg/mL total protein and normal skin tissue homogenate supernatant of 30, 50, and 100 µg/mL total protein, respectively. After cultured for 48 h, the exosomes stimulated with normal protein of 30, 50, and 100 µg/mL and exosomes stimulated with inflammatory protein of 30, 50, and 100 µg/mL were extracted. Normal exosomes, exosomes stimulated with 30 µg/mL normal protein, and exosomes stimulated with 30 µg/mL inflammatory protein were collected, the morphology was observed by transmission electron microscope, the particle size was detected by nanoparticle tracking analyzer, and the expressions of CD9 and CD63 were detected by Western blotting. (3) Twenty one-day-old C57BL/6 mice were taken to isolate the primary passage of fibroblasts (Fbs) and the 3rd passage of Fbs, whose morphology was observed under the inverted phase contrast microscope. The Fbs of 3rd passage were collected to observe the expression of vimentin by cell crawling method combined with immunofluorescence method at culture hour (CH) 2. (4) The Fbs of 3rd passage were divided into control group, normal exosome group, 30, 50, 100 µg/mL normal protein stimulating exosome group, and 30, 50, 100 µg/mL inflammatory protein stimulating exosome group according to the random number table, with 4 wells in each group. Cells in control group received no treatment, and cells in the other 7 groups were respectively added with normal exosomes, exosomes stimulated with normal protein of 30, 50, and 100 µg/mL, and exosomes stimulated with inflammatory protein of 30, 50, and 100 µg/mL prepared in experiment (2). The final mass concentration of exosomes was adjusted to 10 µg/mL. The cell viability was detected by cell count kit 8 at CH 48. (5) Two batches of Fbs in the 3rd passage were divided and treated as those in experiment (4), with 4 wells in each group, and the final mass concentration of exosomes was adjusted to 1 and 10 µg/mL, respectively. The cell mobility was detected by cell scratch test at CH 6, 12, and 24. (6) Two batches of the Fbs of 3rd passage were collected, divided, and treated as those in experiment (4) except with no control group, with 3 wells in each group, and the final mass concentration of exosomes was respectively adjusted to 1 and 10 µg/mL. The mRNA expression levels of transforming growth factor ß(1) (TGF-ß(1)), TGF-ß(3), and α smooth muscle actin (α-SMA) were detected by real-time fluorescent quantitative reverse transcription polymerase chain reaction at CH 48. Data were statistically analyzed with analysis of variance for repeated measurement, one-way analysis of variance, and Bonferroni method. Results: (1) The content of TNF-α in inflammatory wound tissue homogenate supernatant of mice was (116±3) pg/mL, significantly higher than (97±5) pg/mL in normal skin tissue homogenate supernatant at post injury hour 48 (t=3.306, P<0.05). (2) Normal exosomes, exosomes stimulated with 30 µg/mL normal protein, and exosomes stimulated with 30 µg/mL inflammatory protein of hUCMSCs showed the typical saucer-like shape. The particle sizes of the three exosomes of hUCMSCs were 30-150 nm, which were all within the normal particle size range of exosome. Three exosomes of hUCMSCs positively expressed CD9 and CD63. (3) The primary passage of cells were clearly defined and showed protruding spindle shape, irregular polygon shape, or slender strip shape. The morphology of the 3rd and the primary passage of cells is similar. At CH 2, vimentin in cells was positively expressed, and the cells were identified as Fbs. (4) At CH 48, the cell viability was (137.4±2.8)% in 30 µg/mL inflammatory protein stimulating exosome group, obviously higher than 100%, (107.5±2.4)%, (113.3±3.2)%, (104.0±2.0)%, and (101.9±1.5)% in control group, normal exosome group, 30 µg/mL normal protein stimulating exosome group, and 50 and 100 µg/mL inflammatory protein stimulating exosome groups, respectively (P<0.01), and cell viability in 30 µg/mL normal protein stimulating exosome group was obviously higher than that in control group, normal exosome group, and 50 and 100 µg/mL normal protein stimulating exosome groups [(103.4±2.2)% and (102.5±1.4)%], respectively (P<0.01). (5) At CH 6, 12, and 24, the mobility rate of cells in 30 µg/mL inflammatory protein stimulating exosome group was significantly higher than that in control group, normal exosome group, 30 µg/mL normal protein stimulating exosome group, and 50 and 100 µg/mL inflammatory protein stimulating exosome groups, respectively, when the final mass concentrations of exosome was 1 µg/mL (P<0.05) . At CH 12, the mobility rate of cells in 30 µg/mL normal protein stimulating exosome group was obviously higher than that in control group, normal exosome group, and 50 and 100 µg/mL normal protein stimulating exosome groups, respectively, when the final mass concentration of exosome was 1 µg/mL (P<0.05). At CH 6, the mobility rate of cells in 30 µg/mL inflammatory protein stimulating exosome group was significantly higher than that in control group and normal exosome group (P<0.05), and the mobility rate of cells in 30 µg/mL normal protein stimulating exosome group was significantly higher than that in 50 and 100 µg/mL normal protein stimulating exosome groups, respectively, when the final mass concentration of exosome was 10 µg/mL (P<0.05). At CH 12 and 24, the mobility rate of cells in 30 µg/mL inflammatory protein stimulating exosome group was significantly higher than that in control group, normal exosome group, and 50 and 100 µg/mL inflammatory protein stimulating exosome groups (P<0.05), and the mobility rate of cells in 30 µg/mL normal protein stimulating exosome group was significantly higher than that in control group, normal exosome group, and 50 and 100 µg/mL normal protein stimulating exosome groups, respectively, when the final mass concentration of exosome was 10 µg/mL (P<0.05). (6) There were no statistically significant differences in mRNA expression levels of TGF-ß(1), TGF-ß(3), and α-SMA of cells among the 7 groups at CH 48 when the final mass concentration of exosome was 1 µg/mL (F=1.123, 1.537, 1.653, P>0.05). There were no statistically significant differences in mRNA expression levels of TGF-ß(1) and α-SMA of cells among the 7 groups at CH 48 when the final mass concentration of exosome was 10 µg/mL (F=1.487, 1.308, P>0.05), and mRNA expression level of TGF-ß(3) of cells in 50 µg/mL inflammatory protein stimulating exosome group at CH 48 was significantly higher than that in normal exosome group, 50 µg/mL normal protein stimulating exosome group, and 30 and 100 µg/mL inflammatory protein stimulating exosome groups when the final mass concentration of exosome was 10 µg/mL (P<0.05). Conclusions: The pretreatment with inflammatory wound tissue homogenate supernatant of mice has no significant effect on the total protein of hUCMSCs exosomes. The hUCMSCs exosomes stimulated by low concentration inflammatory wound tissue homogenate supernatant can significantly promote the proliferation and migration ability of Fbs. The content of inflammatory mediators in the wound tissue homogenate supernatant during the inflammatory phase is extremely low, which may be the reason that the anti-inflammation and tissue repair paracrine effects of mesenchymal stem cell cannot be effectively started.

[Research progress on surface treatment technology of zirconia implant].

Zirconia material has a color closer to human natural teeth, and has excellent mechanical properties and good biocompatibility, therefore it is expected to become an ideal material for dental implants. In order to improve the osteogenic properties of zirconia implants, a variety of surface treatment techniques emerge in two categories: non-coating method and coating method. Different surface treatment technologies have their advantages and disadvantages. In this paper, the recent progress of zirconia surface treatment technology is reviewed.

[Pepsin and bile acids detection in saliva for the diagnosis of gastroesophageal reflux disease].

Objective: To identify the value of the detection of pepsin and bile acids in saliva for the diagnosis of gastroesophageal reflux disease(GERD). Methods: From January 2018 to June 2019, 104 GERD patients and 43 healthy people in Guangdong Provincial People's Hospital were recruited. The 104 patients of GERD group were divided into four sub-groups, including esophageal symptoms GERD group, extraesophageal symptoms GERD group, anxiety or depression group, non-anxiety and non-depression group. Saliva was collected on waking in morning and 2 h after finishing lunch. The concentration of the total pepsin(TPP) and total bile acids(TBA) from saliva was detected by ELISA method. Receiver operating characteristics analysis was used to identify the sensitivity and specificity of the saliva pepsin and bile acids detection. Results: The concentration of TPP in morning waking samples and postprandial samples in the GERD group was 27.1(9.7,50.3) µg/L and 32.4(14.0,58.7) µg/L, the concentration of TBA in postprandial samples was (18.4±2.3)µmol/L, and these levels were significantly higher than that of the control group [7.0(5.1, 9.1) µg/L, 7.4(5.2, 9.4) µg/L, (12.6±5.0)µmol/L](P<0.01). The concentration of TBA in morning waking samples had no significant difference between these two groups(P>0.05). The concentration of TPP and TBA had no significant difference among the four GERD sub-groups(P>0.05).Pepsin in postprandial saliva samples had moderate diagnostic value for GERD, when the saliva pepsin concentration in postprandial samples was higher than 41.33 µg/L, it had a sensitivity of 82.8% and a specificity of 73.3%. The bile acids in saliva had no significant diagnostic value for GERD. Conclusions: Pepsin detection in saliva has a high level of sensitivity and specificity for diagnosing GERD. However, bile acids in saliva has no significant diagnostic value for GERD.

[Advances in the research of effects of epithelial-mesenchymal interaction on wound healing and scar formation].

Wound healing is a dynamic process which involves interaction of various types of cells, cytokines, and extracellular matrix. Among them, epithelial cells and mesenchymal cells are the key components which involve in wound healing and scar formation. Related scholars had done a great number of studies about the functions of epithelial cells and fibroblasts(Fbs) in wound healing and scar formation. The results showed that under the stimulation of complex microenvironment, epithelial cells would lose their epithelial characteristics and acquire the typical characteristics and migration ability of mesenchymal cells. At the same time, with the complex changes of cell structure and cell behavior, they would participate in the process of tissue wound repair, including normal or fibrotic repair, by covering the wound with migration. Fbs are the key cells for the wound fibrotic repair, and play important roles in the process of wound healing, including excessive wound healing or delayed wound healing. In the recent years, the researchers realized that the cross-talk between epithelial cells and Fbs in wound healing, which is referred to as epithelial-mesenchymal interaction, significantly changes the biological behaviors of these two cell types, which affects the dermal remodeling and re-epithelialization quality of wound. Epithelial-mesenchymal interaction plays an important role in skin morphogenesis during embryonic development and maintaining the structural integrity of adult skin. In the process of re-epithelialization, Fbs could promote the proliferation and migration of keratinocytes, meanwhile keratinocytes would receive the signals from Fbs to reconstruct functional epithelium, which has become a hot topic in the field of wound healing at present. In this paper, a comprehensive analysis of the literature on the role of epithelial-mesenchymal interaction in wound healing and scar formation at home and abroad in recent years is presented for the reference of relevant scholars.

[Influence of human amniotic mesenchymal stem cells on macrophage phenotypes and inflammatory factors in full-thickness skin wounds of mice].

Objective: To explore the influence of human amniotic mesenchymal stem cells (hAMSCs) on the in vivo and in vitro regulation of macrophage phenotypes and inflammatory factors associated with wound healing of full-thickness skin wounds in mice. Methods: Fresh amniotic membrane discarded from full-term delivery by 5 healthy pregnant women in the Department of Obstetrics and Gynecology of the Affiliated Hospital of Zunyi Medical University was used for the isolation and culture of hAMSCs by enzyme digestion method. The third passage of cells was used for identification of adipogenic and osteogenic differentiation. The fourth passage of cells was used for identification of hAMSCs surface markers. Ten C57BL/6 mice (all male, aged 6 to 8 weeks, the same gender and age below) were selected for extracting mouse peritoneal macrophages by intraperitoneal lavage, and M1-type macrophages were induced by Dulbecco's modified eagle medium (DMEM) medium containing interferon-γ. The M1-type macrophages were divided into hAMSCs+ macrophage group and macrophage alone group. Then 1×10(4) hAMSCs/per well of fourth passage were added to macrophage in hAMSCs+ macrophage group and cultured in 2 mL DMEM medium for routine culture. In macrophage alone group, each well was only added with 2 mL DMEM medium for routine culture. On day 1 and 7 in culture, the content of interleukin-12 (IL-12), arginase 1, and IL-10 in the cell culture supernatant of the 2 groups were detected by enzyme-linked immunosorbent assay with sample number of 6/per group. (2) Full-thickness skin wound model was reproduced in the back of 56 C57BL/6 mice, which were divided into hAMSCs group and phosphate buffer solution (PBS) group using the random number table, with 28 mice in each group. Mice in hAMSCs group were subcutaneously injected with 100 µL of cell suspension containing 1×10(7) hAMSCs per mL in PBS suspension along the wound edge. While mice in PBS group were only subcutaneously injected with 100 µL PBS along the wound edge. On post injection day (PID) 1, 3, 7, and 14, 7 mice in the two groups were sacrificed respectively. Histopathological observation was performed with hematoxylin-eosin staining. The expressions of macrophage surface markers [CD68 and inducible nitric oxide synthase (iNOS) double positive cells and CD68 and arginase 1 double positive] in the wounds were detected by immunofluorescent staining. The mRNA expressions of IL-10, macrophage inflammatory protein 1α (MIP-1α), and MIP-2 in the wounds were detected by real-time fluorescent quantitative reverse transcription polymerase chain reaction. Data were statistically analyzed with analysis of variance for factorial design, t test, and Bonferroni correction. Results: (1) On day 1 in culture, the content of IL-12 and arginase 1 in the cell culture supernatant of the two groups were similar (t=0.448, 0.536, P>0.05), and the content of IL-10 in the cell culture supernatant of hAMSCs+ macrophage group was significantly lower than that in macrophage alone group (t=14.722, P<0.01). On day 7 in culture, the content of IL-12 in the cell culture supernatant of hAMSCs+ macrophage group was significantly lower than that in macrophage alone group (t=13.226, P<0.01), and the content of arginase 1 and IL-10 was significantly higher than that in macrophage alone group (t=30.172, 31.406, P<0.01). (2) On PID 1, a large number of inflammatory cells infiltration were observed in the skin wounds of both groups. On PID 3, the inflammatory cells infiltration in the skin wounds increased in both groups, and the inflammatory cells infiltration in hAMSCs group was less than that in the PBS group. On PID 7, the inflammatory cells infiltration in the wounds decreased in both groups, and the inflammatory cells infiltration in hAMSCs group was less than that in the PBS group. On PID 14, no obvious inflammatory cells infiltration was observed in the wounds in the two groups. (3) On PID 1 and 14, the percentages of CD68 and iNOS double positive cells and CD68 and arginase 1 double positive cells in the wounds were similar in the two groups (t(1 d)=0.134, 0.693, t(14 d)=1.146, 2.585, P>0.05). On PID 3 and 7, the percentages of CD68 and iNOS double positive cells in the wounds in hAMSCs group were significantly lower than those of PBS group (t=6.396, 4.787, P<0.01), while the percentages of CD68 and arginase 1 double positive cells were significantly higher than those of PBS group (t=3.928, 4.473, P<0.01). (4) On PID 1, the mRNA expressions of IL-10 in the wounds of mice in the two groups were similar (t=2.005, P>0.05). On PID 3, 7, and 14, the mRNA expressions of IL-10 in the wounds of mice in hAMSCs group were significantly higher than those of PBS group (t=7.758, 124.355, 80.823, P<0.01). On PID 1, 3, 7, and 14, the mRNA expressions of MIP-1α and MIP-2 in the wounds of mice in hAMSCs group (0.341±0.212, 0.648±0.004, 0.611±0.106, 0.763±0.049, 1.377±0.099, 1.841±0.042, 1.181±0.035, 0.553±0.028) were significantly lower than those of PBS group (3.853±0.035, 6.914±0.163, 3.648±0.113, 2.250±0.046, 11.119±0.495, 8.634±0.092, 5.722±0.021, 4.862±0.036, t=43.198, 101.904, 51.845, 58.231, 51.074, 177.501, 291.752, 251.614, P<0.01). Conclusions: hAMSCs demonstrates biological effects of promoting the transformation of M1-type macrophages into M2-type macrophages in full-thickness skin wounds of mice. They can up-regulate the expression of anti-inflammatory and anti-fibrotic factor IL-10, and down-regulate the expression of important inflammation mediated factors MIP-1α and MIP-2.

[Analysis of early renal damage markers under different renal ultrasonic manifestations in gout patients].

Objective: To observe the relationship between early renal damage markers and renal ultrasonic manifestations in gout patients, and whether early renal damage is reversible after uric acid-reducing treatment. Methods: The gout patients from the Department of Rheumatology of Beijing Haidian Hospital and Peking University People's Hospital between July 2016 and December 2017 were recruited in this study. According to the results of renal ultrasonography, the patients were divided into the following three groups. Group A was normal. Group B was punctate crystallization. Group C was renal calculi. Each group included 30 patients. The patients in group B and group C who could insist on regular uric acid-reducing treatment for one year were selected. The levels of urinary RBP, ß(2)-MG and NAG were measured in different groups and one year before and after uric acid-reducing treatment. Results: The urinary concentration of ß(2)-MG in group A, group B and group C were (128.59±107.32), (316.08±207.41) and (311.25±162.85)mg/L, respectively. There were significant differences among the three groups (P<0.001). The urinary concentration of NAG were (13.41±5.12)U/L,(17.88±6.19)U/L and (18.48±9.84)U/L, respectively. There were differences among the three groups (P<0.01).There was no significant difference in urinary RBP concentration among the three groups (P=0.188). After one year of uric acid-reducing treatment, the levels of urinary RBP, ß(2)-MG and NAG were lower than that before treatment. There were significant differences before and after treatment in each group (P<0.05). Compared with group C, the levels of urinary ß(2)-MG and NAG were decreased in group B after uric acid-reducing treatment (all P<0.05). Conclusions: Renal ultrasonography is helpful for the diagnosis of early renal damage in gout patients. Early renal damage markers in gout patients decreased after uric acid-reducing treatment, suggesting that early renal damage can be reversible if early diagnosis and timely treatment can be made in gout patients.

[Cost-effectiveness analysis of rabies immunization strategy based on dynamic-decision tree model].

Objective: To evaluate the cost-utility of different immunization strategies for rabies in China, and to provide a reference for determining the optimal immunization strategy. Methods: The system dynamics model was used to simulate the epidemic of canine rabies and a decision tree model was conducted to analysis different immune strategies. Relevant probabilities were obtained through literature search and on-site investigation. Sensitivity analysis was used to explore the important influenced factors. Results: At baseline, from a social perspective, 70% vaccination of dogs was the optimal strategy compared to current vaccination strategy (43% vaccination in dogs, human category-Ⅱ exposure vaccination/category-Ⅲ exposure vaccination combined with RIG). The total cost was 14 084 354 CNY, and the total utility value was 22 078 616.23 QALYs, and the incremental cost-utility ratio was-62 148 147 CNY/QALY; if human vaccination was considered, 55% vaccination of dogs combined with strategy one was the optimal strategy, its incremental cost-utility ratio was-444 620 557 CNY/QALY. The probability that an injured dog carries rabies virus was the most sensitive parameter. When it was greater than 0.005 03, strategy four was the optimal strategy. When it was less than 82/100 000, strategy one was the optimal strategy; when it was between 82/100 000 and 120/100 000, strategy two was the optimal strategy; when it was between 120/100 000 and 503/100 000, strategy two was the optimal strategy. Conclusion: It was conducive to increase the vaccination coverage of canine for the prevention and control of rabies.

[Clinical effects of free superficial femoral artery femoral triangle perforator flap in the repair of skin and soft tissue defects in extremities].

Objective: To investigate the clinical effects of free superficial femoral artery femoral triangle perforator flap in the repair of skin and soft tissue defects in extremities. Methods: From January 2016 to November 2017, 14 patients (9 males and 5 females, aged 19 to 54 years) with skin and soft tissue defects in extremities accompanied with tendon and bone exposure were admitted to our unit. The size of skin and soft tissue defects after debridement ranged from 7 cm×3 cm to 10 cm×7 cm. The defects were repaired with free superficial femoral artery femoral triangle perforator flaps, with size ranging from 13.0 cm×2.0 cm to 20.0 cm×4.5 cm. The medial femoral cutaneous nerve was applied to the flap. The perforator flap was grafted onto the medial femoral cutaneous nerve in 6 patients. The donor sites were sutured directly. The survival of flaps and the follow-up of patients were observed. Results: All flaps of 14 patients survived successfully. The recipient sites and donor sites were healed completely in 13 patients, and 1 patient with partial skin necrosis at the edge of flap was healed after treatment. All patients were followed up for 6 months to 1 year after the operation. The flaps were in good shape, with nearly normal color and soft texture and no cicatrix contracture deformity. The flaps recovered protective sense in 6 patients who had medial femoral cutaneous nerve grafting, and the sensory recovery of the flap was slightly worse in the remaining 8 patients. There was no significant complications on the appearance and walking of the donor thigh in 14 patients, only a linear scar was left on the inner thigh, and no numbness was felt in the donor sites of patients. Conclusions: The free superficial femoral artery femoral triangle perforator flap is an ideal therapy for repairing skin and soft tissue defects in extremities.

[Effect of perforator flap of the proper digital artery of the ulnar or radial side of finger in the treatment of webbed scar contracture of the same finger in child].

Objective: To explore the effect of the perforator flap of the proper digital artery on the ulnar or radial side of the finger in the treatment of webbed scar contracture of the same finger in child. Methods: From January 2012 to January 2016, 26 children who were treated with dressing change after burn of finger and then had webbed scar contracture along with growth and development were hospitalized in our unit, involving a total of 50 fingers. There were 14 males and 12 females among the children aged from 2 to 14 years. After the scar was dissected and released, the wound area ranged from 1.6 cm×1.0 cm to 5.0 cm×2.6 cm. The perforator flap of the proper digital artery of the ulnar or radial side of the same finger was used to repair the wound. The flap area ranged from 1.8 cm×1.0 cm to 4.6 cm×1.8 cm. The donor sites were sutured directly. The residual wounds in donor and recipient sites were repaired by full-thickness skin graft collected from inguinal area/adjacent area or adjacent perforator flap. The postoperative development and function of the fingers were followed up and observed. The range of motion of the fingers was evaluated according to the Chinese Medical Association Hand Surgery Society's upper limb functional evaluation trial standard, the Kantor Scar Cosmesis Assessment and Rating Scale was used to score the scar of finger, and the latest data were recorded. Results: The flaps and skin grafts survived successfully after operation. The patients were followed up for 6 to 24 months. The perforator flaps of the proper digital artery on the ulnar or radial side of the finger survived well at the latest follow-up, with good color and texture and a two-point discrimination distance of 9 to 12 mm. There was no contracture of the fingers, a little pigmentation in the skin graft area, no flexion deformity of the fingers, no lateral bending of the fingers to the flap-harvesting side, and no scar contracture at the webs of the fingers. Compared with that of healthy side, the development of finger was not obviously abnormal. The range of motion of the fingers was excellent in 38 fingers and good in 12 fingers, and the scar score of the fingers was 2-3 points in 31 fingers, 4-7 points in 15 fingers, and 8-10 points in 4 fingers. Conclusions: The efficacy of perforator flap of the proper digital artery of the ulnar or radial side of finger in the treatment of the webbed scar contracture of the same finger in child is reliable, with high postoperative survival rate of the flap, better color and texture, and fewer complications, which can avoid the risk of re-contracture of the finger in a short period after operation, and does not affect the growth and development of the finger.

[Clinical effects of superior gluteal artery perforator island flap in repair of sacral pressure ulcer].

Objective: To explore the clinical effects of superior gluteal artery perforator island flap in repair of sacral pressure ulcer. Methods: From May 2012 to May 2017, 20 patients with sacral pressure ulcers (14 males and 6 females, aged 27 to 67 years) were admitted to our department. According to the consensus staging system of National Pressure Ulcer Advisory Panel in 2016, 6 cases were in 3 stages, 14 cases were in 4 stages, with the area of pressure ulcers ranging from 5.0 cm×4.0 cm to 10.0 cm×8.0 cm. After debridement and vacuum sealing drainage, the superior gluteal artery perforator island flaps were used to repair the pressure wounds, with the area of flaps ranging from 6 cm×5 cm to 13 cm×8 cm. The donor sites were sutured directly. The survival of flaps after operation, the healing of wounds, and the follow-up of patients were observed. Results: After surgery, flaps of 20 patients survived well without reoperation. The length of hospital stay of patients was 20 to 40 days, with an average of 25 days. Eighteen patients were followed up for 6 to 24 months, with an average of 12.2 months. The flaps were in good shape and elastic recovery. There were no complications such as seroma or hematoma in the donor sites. Both the patients and family members expressed satisfaction with the shape and texture of the flap and shape of hip. Conclusions: The superior gluteal artery perforator island flap is reliable in blood supply and easy to rotate. The flap can carry a little muscle to increase the anti-infective ability. Moreover, the donor site can be directly sutured with slight damage. Thus, it is one of the good methods for repairing sacral pressure ulcers.

[The value of the planned neoadjuvant radiotherapy or chemoradiotherapy for the non-radical resection of esophageal squamous cell carcinoma].

Objective: The role of planned neoadjuvant radiotherapy or chemoradiotherapy in the non-radical resection of esophageal squamous cell carcinoma was unclear. The study aimed to evaluate their therapeutic effect and analyze the prognostic factors. Methods: We retrospectively analyzed the clinical data of locally advanced esophageal squamous cell carcinoma who received neoadjuvant radio therapy (33 patients) and concurrent chemoradiotherapy (119 patients) from January 2004 to December 2016 in our single-institution database.The survival rates were calculated by Kaplan-Meier method. The prognostic factors were analyzed by using Log rank test and Cox proportional hazards model. Results: The median follow-up was 29.8 months. One hundred and one patients survived more than 3 years. The rates of overall survival (OS) and disease-free survival (DFS) at 3 years were 63.9% and 55.6%, respectively.The rates of complete, partial and minimal pathological response of the primary tumor were 50.3%, 38.4%, 11.3%, the corresponding 3-year OS were 75.5%, 57.4%, 27.3% (P<0.001) and 3-year DFS were 72.0%, 44.7%, 17.6% (P<0.001), respectively.The postoperative lymph node metastasis rate was 27.0%. The 3-year OS and DFS of the lymph node positive group was 45.6% and 32.8%, significantly lower than 70.8% and 63.7% of the negative group (both P<0.001). The 3-year OS and DFS of pathologic stage Ⅰ, Ⅱ, ⅢA, ⅢB and Ⅵ A were 76.2%, 57.4%, 64.7%, 35.0%, 33.3% (P<0.001) and 70.1%, 49.3%, 41.2%, 22.1%, 33.3% (P<0.001), respectively.The operation-related mortality was 3.3%. Multivariate analysis showed that chest pain, postoperative respiratory failure, pathological differentiation, more than 15 lymph node dissection and ypTNM stage were the independent prognostic factors of OS (P<0.05 for all). Conclusions: The planned neoadjuvant radiotherapy or chemoradiotherapy for the non-radical resection of advanced esophageal squamous cell carcinoma could result in favorable survival. The chest pain, postoperative respiratory failure, pathological differentiation, the number of lymph node resection and ypTNM stage are the independent prognostic factors of the prognosis of these patients.

[Effects of combined transplantation of rat Schwann cells and fibroblasts on nerve regeneration of denervated perforator flaps in rats and the mechanism].

Objective: To explore the effects of combined transplantation of the rat Schwann cells and fibroblasts (Fbs) on the nerve regeneration of denervated perforator flaps in rats and the mechanism. Methods: (1) Fbs were isolated from the trunk of 2 Sprague-Dawley (SD) rats embryos of 14-16 days' pregnancy and cultured, and the morphology of the cells was observed. The third passage of cells were used for subsequent experiments. The protein expressions of fibronectin and Ephrin-B2 were observed by immunohistochemical method. The mRNA expression of Ephrin-B2 was detected by real-time fluorescent quantitative reverse transcription polymerase chain reaction (n=3). (2) Schwann cells were isolated from the bilateral sciatic nerves and brachial plexus nerves of 45 SD rats born for 1-3 days and cultured, and the morphology of the cells was observed. The third passage of cells were used for subsequent experiments. The rate of S100 positive cells was detected by immunofluorescence method and flow cytometer, with sample numbers of 9 and 3 respectively. (3) In Dulbecco's modified Eagle medium (DMEM) high glucose medium, 1 mL Fbs and 1 mL Schwann cells both in the concentration of 1×10(5) cells/mL were co-cultured as Schwann cells+ Fbs co-culture group, and 2 mL Schwann cells in the concentration of 1×10(5) cells/mL were cultured alone as Schwann cells alone culture group, with 5 wells in each group. The clusters of Schwann cells in the two groups were observed and counted under inverted phase contrast microscope at post culture hour (PCH) 6 and 24 respectively. The clusters of Schwann cells in Schwann cells+ Fbs co-culture group were observed by immunofluorescence method at PCH 24 too. The protein expressions of EphB2, Sox2, and N-cadherin in Schwann cells of two groups at PCH 24 were detected by Western blotting (n=20). (4) Totally 100 8-week-old male SD rats were selected, and an in situ replanted peritoneal denervated perforator flap was made in each rat. According to the random number table, the rats were divided into simple flap group, Fbs alone transplantation group, Schwann cells alone transplantation group, Schwann cells+ Fbs co-transplantation group, with 25 rats in each group. Flaps of rats in Fbs alone transplantation group and Schwann cells alone transplantation group were injected with 0.4 mL Fb and 0.4 mL Schwann cells respectively (2×10(6) cells each). Flaps of rats in Schwann cells+ Fbs co-transplantation group were injected with 0.4 mL Fbs and Schwann cells mixed cells (totally 2×10(6) cells, cell number ratio: 1∶1), and flaps of rats of simple flap group were injected with the same volume of DMEM high glucose medium. On post injection day (PID) 2, 5, 7, 9, and 14, 5 rats in each group were selected respectively according to the random number table. The flap tissue was collected, and the number, diameter, and arrangement of regenerated nerves were observed by immunofluorescence method. Data were processed with completely random designed t test, analysis of variance for repeated measurement, t test, and Bonferroni correction. Results: (1) The third passage of cells isolated and cultured from the rat embryo trunks were uniform in size and shape, long spindle-shaped, with a large proportion of nuclei. Strong positive expressions of fibronectin and Ephrin-B2 protein in cells were observed, and the mRNA expression of Ephrin-B2 was 0.004 1±0.000 8. The cells were identified as Fbs. (2) After 5 days of culture, the primary cells isolated from the sciatic nerves and brachial plexus nerves of neonatal rats were elongated in cell bodies and grew in nest, fence, or vortex-like shape. The third passage of cells were detected by immunofluorescence method and flow cytometer, and the corresponding S100 positive cell rates were (95.9±1.0)% and (95.8±1.1)% respectively. The cells were identified as Schwann cells. (3) At PCH 6 and 24, the cluster numbers of Schwann cells in Schwann cells+ Fbs co-culture group were significantly higher than those of Schwann cells alone culture group (t=6.500, 10.614, P<0.01). At PCH 24, the Schwann cells in Schwann cells+ Fbs co-culture group aggregated into clusters, Fbs dispersed around the Schwann cell clusters, and the protein expressions of EphB2, N-cadherin, and Sox2 in Schwann cells were significantly higher than those in Schwann cells alone culture group (t=2.975, 19.717, 11.159, P<0.05 or P<0.01). (4) On PID 2, a small number of scattered, disordered, short, and thin nerve fibers were observed in the flap tissue of rats in the four groups. From PID 5 to 14, the number of nerve fibers in the flap tissue of rats of Schwann cells+ Fbs co-transplantation group increased gradually, and the nerve fibers were with long diameter and arranged orderly. The number of nerve fibers in the flap tissue of rats of Schwann cells alone transplantation group increased, but the nerve fibers were with short diameter and arranged disorderly, and the number was smaller than that of Schwann cells+ Fbs co-transplantation group. In simple flap group and Fbs alone transplantation group, the nerve fibers in the flap tissue of rats gradually degenerated with gradually decreased number or even disappeared. Conclusions: The combined transplantation of Fbs and Schwann cells in rats can regulate Schwann cells migration and clustering by activating Ephrin/Eph-Sox2-N-cadherin signaling pathway, thus promoting the orderly nerve regeneration of denervated perforator flaps in rats.