Efficient gene editing in a medaka (Oryzias latipes) cell line and embryos by SpCas9/tRNA-gRNA / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Generation of mutants with clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) is commonly carried out in fish species by co-injecting a mixture of Cas9 messenger RNA (mRNA) or protein and transcribed guide RNA (gRNA). However, the appropriate expression system to produce functional gRNAs in fish embryos and cells is rarely present. In this study, we employed a poly-transfer RNA (tRNA)-gRNA (PTG) system driven by cytomegalovirus (CMV) promoter to target the medaka (Oryzias latipes) endogenous gene tyrosinase(tyr) or paired box 6.1 (pax6.1) and illustrated its function in a medaka cell line and embryos. The PTG system was combined with the CRISPR/Cas9 system under high levels of promoter to successfully induce gene editing in medaka. This is a valuable step forward in potential application of the CRISPR/Cas9 system in medaka and other teleosts.

Preliminary effect observation on the application of micro-negative pressure in children with small-area deep partial-thickness burn / 中华烧伤杂志

Objective@#To preliminarily observe the effects of application of micro-negative pressure in children with small-area deep partial-thickness burn.@*Methods@#From January 2016 to August 2018, 64 children with small-area deep partial-thickness burn who were admitted to the Department of Burn Surgery of the First Affiliated Hospital of Naval Medical University were recruited in this prospective randomized controlled study. According to the random number table, they were divided into negative pressure group [18 boys and 14 girls, aged (3.9±1.6) years with total burn area of (5.5±2.2)% total body surface area (TBSA)] and conventional group [20 boys and 12 girls, aged (3.8±1.7) years with total burn area of (5.8±1.6)% TBSA], with 32 patients in each group. After admission, simple debridement was performed in the patients of 2 groups. After that, the children in negative pressure group were treated with micro-negative pressure with negative pressure material replaced every 3 to 5 days. Children in conventional group were treated with silver sulfadiazine cream with dressing change every other day. On post injury day (PID) 14 and 21, general wound observation was performed, the wound healing rate was calculated, the exudates from the wounds were cultured and the positive detection rate was calculated. The number of patients requiring surgical skin grafting was recorded and the rate of surgical skin grafting was calculated, and the complete wound healing time was recorded in the patients of 2 groups. Scar formation was evaluated by the Vancouver Scar Scale (VSS) in 3, 6, and 12 months after wound healing. Data were processed with chi-square test, t test, Bonferroni correction, and analysis of variance for repeated measurement.@*Results@#(1) On PID 14, all the necrotic tissue in the wounds of patients in negative pressure group was removed, with few exudates, and most of the wounds had been epithelialized; most of necrotic tissue in the wounds of patients in conventional group was removed, with more exudates and smaller wound healing area than those in negative pressure group. On PID 21, most of the wounds of patients in negative pressure group were healed, and the exudates were rare, while the wound healing area of patients in conventional group was significantly smaller than that in negative pressure group with more exudates. (2) On PID 14 and 21, the wound healing rates [(49.8±3.3)% and (95.8±2.4)%] of patients in negative pressure group were significantly higher than those in conventional group [(40.0±3.2)% and (75.3±2.5)%, t=11.899, 33.461, P<0.01]. (3) On PID 14 and 21, the positive detection rates of wound bacteria of patients in negative pressure group were significantly lower than those in conventional group (χ2=6.275, 5.741, P<0.05). (4) The rate of surgical skin grafting of patients in negative pressure group was significantly lower than that in conventional group (χ2=5.333, P<0.05). (5) The complete wound healing time of patients in negative pressure group [(23.9±2.3) d] was significantly shorter than that in conventional group [(27.9±1.8) d, t=-7.806, P<0.01]. (6) In 3, 6, and 12 months after wound healing, the VSS scores [(6.9±1.8), (5.6±1.4), (3.4±1.5) points] of patients in negative pressure group were significantly lower than those in conventional group [(9.0±1.5), (7.4±2.0), (5.7±1.6) points, t=-4.987, -4.127, -5.988, P<0.01].@*Conclusions@#In comparison with routine dressing change, the treatment of application of micro-negative pressure in children with small-area deep partial-thickness burn can significantly improve the wound healing rate and rate of surgical skin grafting, decrease the wound infection rate, shorten the wound healing time, and improve the wound healing quality.

Debridement combined with vacuum sealing drainage in the treatment of severe infection in abdominal wall due to allogeneic umbilical cord embedded in abdominal wall for immunotherapy / 中华烧伤杂志

Objective@#To explore the effect of debridement combined with vacuum sealing drainage (VSD) on the treatment of severe infection in abdominal wall due to allogeneic umbilical cord embedded in abdominal wall for immunotherapy.@*Methods@#From January 2015 to December 2016, 12 patients with severe infection in abdominal wall due to allogeneic umbilical cord embedded in abdominal wall for immunotherapy were admitted to our department. They were conducted with systemic anti-infective treatment, local debridement, and VSD. The wounds were continuously washed for 3 to 5 days after the VSD device installed, with negative pressure value from -16.0 to -12.0 kPa. The VSD device was removed 5 to 7 days later. Continue wound dressing by aseptic ribbon gauze was stuffed in the cavity, and the incision was sutured after the granulation tissue grew well in the cavity.@*Results@#In all patients, allogeneic umbilical cords were completely removed and abdominal infection was cured. The wounds healed well, the sensory function of abdominal was normal, and the activity was not restricted. All the patients were followed up for 3 to 6 months with no reinfection or incisional hernia.@*Conclusions@#Embeding the whole allogeneic umbilical cord in abdominal wall for immunotherapy can lead to severe infection in abdominal wall. Abdominal infection can be cured by debridement combined with VSD with good clinical results.

Effects of caspase-1 inhibitor VX765 on cold-restraint stress-induced acute gastric ulcer in mice / 中华烧伤杂志

Objective@#To investigate the protective effects of caspase-1 inhibitor VX765 on gastric mucosa of mice with cold-restraint stress-induced acute gastric ulcer.@*Methods@#Twenty-four specific pathogen free male C57BL/6J mice were divided into normal control group (NC), cold restrain group (CR), VX765 pre-treatment+ cold restrain group (VCR), and rabeprazole pre-treatment+ cold restrain group (RCR) according to the random number table, with 6 mice in each group. Mice in group NC were injected intraperitoneally with solution of 10 mL/kg dimethylsulfoxide (DMSO) and phosphate buffer solution (PBS). Mice in group CR were inflicted with acute gastric ulcer induced by cold-restraint stress 30 minutes after intraperitoneal injection of solution of DMSO and PBS. Mice in groups VCR and RCR were inflicted with acute gastric ulcer as above 30 minutes after intraperitoneal injection of solution of DMSO and PBS with dose of 12.5 μmol/kg containing 10 mg VX765 and 40 mg/kg containing 20 mg rabeprazole, respectively. Four hour after cold-restraint stress, serum content of tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6)was determined by enzyme-linked immunosorbent assay. Gross condition of gastric tissue was observed. Ulcer index was evaluated. Pathological change of gastric tissue was observed with HE staining. The relative expression of IL-1β, IL-18, and cleaved-caspase-1 in gastric tissue were detected by Western blotting. Mice in group NC were detected as above at the same time point. Data were processed with one-way analysis of variance and Bonferroni test.@*Results@#The serum content of TNF-α and IL-6 and the relative expression of cleaved-caspase-1, IL-1β, and IL-18 in gastric tissue of mice in group NC were significantly lower than those in group CR (with P values below 0.01). The content of the above-mentioned inflammatory indexes in serum and gastric tissue of mice in group VCR was significantly lower than that in group CR (with P values below 0.01). There were no statistically significant differences in content of the above-mentioned inflammatory indexes in serum and gastric tissue of mice between groups RCR and CR (with P values above 0.05). The content of the above-mentioned inflammatory indexes in serum and gastric tissue of mice in group VCR was significantly lower than that in group RCR (with P values below 0.01). Surface of gastric mucosa was smooth and morphology of mucosal cells was normal with clear structure of mice in group NC. Multiple hemorrhage of gastric mucosa, disorderly arrangement of mucosal cells, and large number of inflammatory cell infiltration around necrotic tissue were observed in mice of group CR. Decreased number of gastric mucosa bleeding, intact mucosal structure, and small amount of inflammatory cell infiltration around necrotic tissue were observed in mice of groups VCR and RCR. The ulcer indexes of mice in groups NC, CR, VCR, and RCR were 0, 18.7±1.1, 6.3±1.5, and 8.2±1.3, respectively. The ulcer index of mice in group NC was significantly lower than that in the other 3 groups (with P values below 0.05). The ulcer indexes of mice in groups VCR and RCR were close (P>0.05), which were significantly lower than ulcer index of mice in group CR (with P values below 0.05).@*Conclusions@#VX765 can effectively inhibit the activation of caspase-1, reduce production of inflammatory factor, and alleviate inflammatory response, which have protective effects on gastric mucosa of mice with cold-restraint stress-induced acute gastric ulcer.

Advances in preparation and clinical application of amniotic membrane graft / 中华烧伤杂志

Amniotic membrane has been used as wound dressing for more than 100 years. With the development of the preservation and preparation techniques, amniotic membrane is widely used in ophthalmology, burns, plastic surgery, dentistry, and neurosurgery. In recent years, as more and more amniotic membrane is used in chronic wounds, it will be a new treatment method for wounds. This paper is a brief review about advances in preparation and clinical application of amniotic membrane graft.

Comparison of the effects of different doses of methylprednisolone therapy for children with severe hand,foot and mouth disease / 中国基层医药

Objective To compare the clinical effects of different doses of methylprednisolone therapy for children with severe hand,foot and mouth disease (HFMD).Methods According to different dosage methods, 240 children with severe HFMD were divided into large dose group,medium dose group and small dose group,80 cases in each group.The three groups were given different doses of methylprednisolone infusion on the basis of conventional treatment:large dose group(5 ~10mg·kg· -1 d -1 ),medium dose group(3 ~5 mg·kg· -1 d -1 ),small dose group (1 ~2mg · kg · -1 d -1 ).Results The time of fever sustaining,panic ease,mechanical ventilation,duration of hypertension and heart rate recovery of the medium -dose group were (47.93 ±4.72)h,(45.54 ±2.42)h,(51.43 ± 6.85)h,(53.66 ±7.62)h,(52.45 ±7.84)h,which were significantly shorter than those of the small -dose and large -dose group(all P 0.05 ). Conclusion Medium dose of methylprednisolone in the treatment of children with severe HFMD has significant effect and less adverse reactions,which is worthy of promotion.