Objective To construct a RhoA-siRNA expression vector and determine its role on the malig-nant behavior of HepG2 cells.Methods A RhoA-siRNA DNA fragment was synthesized and cloned into the expression vector of pGenesil-1.The constructed Rhon-siRNA DNA plasmid was stably transfected into HerG2 cells by lipofectamine,and then HepG2 cells were divided into the HepG2/RhoA-siRNA group (HepG2 cells were transfected with pGenesil-1-RhoA-siRNA),HepG2/control group(HepG2 cells were transfected with control plasmid) and HepG2 group (without plasmid transfection).The inbibitory effect of RhoA-siRNA on RhoA protein expression was shown by Western blot.The proliferation,migration,growth potentiality and cell cycle of transfected HepG2 cells were evaluated by MTT assay,wounded healing,the plate cloning formation test and flow cytometry,respectively.All data were analyzed by one-way analysis of variance (ANOVA) and chi-square test.Results The expression of RhoA protein in the HepG2/RhoA-siRNA group was,significantly decreased compared with that in the other two groups (F=178.19,P<0.05).Scratched cells were healed within 48 hours in the HepG2/control group and HepG2 group,but not in the HepG2/RhoA-siRNA group.The clone formation rates in the HepG2/RhoA-siRNA group,HepG2 group and HepG2/control group were 39%±3%,67%±5%and 70%±6%,respectively,with a significant difference among the three groups(χ2=33.34,38.69,P<0.05).Flow cytometry showed that the number of cells transfected with RhoA-siRNA was highest in the G0/G1 phase and lowest in the S phase(F=70.46,76.57.P<0.05).Conclusion The RhoA-siRNA expression vector can effectively suppress the proliferation and migration of HepG2 cells,which may provide a novel gene therapy for hepatocellular carcinoma.

Objective: To observe the effects of Na⁺ /H⁺ exchanger 1 (NHE1) inhibitor on intestinal injury of rats with burn sepsis, and to explore the possible mechanism preliminarily. Methods: Ninety SD rats were divided into control group, pure sepsis group, and NHE1 inhibitor group according to the random number table, with 30 rats in each group. Full-thickness scald (hereinafter referred to as burn) model with 20% total body surface area were reproduced on the back of rats in pure sepsis and NHE1 inhibitor groups, and then 50 μL liquid of Pseudomonas aeruginosa ATCC 27853 (2×10⁵ colony forming unit/mL) were injected into the center of wounds on the back. Rats in NHE1 inhibitor group were intraperitoneally injected with 0.1 mmol/L NHE1 inhibitor cariporide (0.4 mg/kg) rapidly after the successful establishment of burn sepsis model, while rats in pure sepsis group were injected with the same volume of normal saline. Except for not being made burn wounds nor receiving bacterination, rats in control group were treated the same as those in pure sepsis group. Rats with burn sepsis in each group were laparotomized and injected with 200 mL fluorescein isothiocyanate (FITC)-dextran in the concentration of 0.1 mol/L in terminal ileum at 12 hours post injury, and their left ventricular blood and terminal ileum were collected 30 minutes later. The serum content of FITC-dextran was detected with fluorescence spectrophotometer (n=10); the morphology of intestinal tissue was observed with HE staining (n=10); the content of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) in serum and intestinal tissue was determined with enzyme-linked immunosorbent assay (n=20); the activity of myeloperoxidase (MPO) in serum and intestinal tissue was detected with colorimetric method (n=20); the protein expression of nuclear factor-kappa B-p65 (NF-κB-p65) and phosphorylation levels of mitogen-activated protein kinase (MAPK) signal pathway related proteins p38MAPK, extracellular signal-regulated kinase 1/2 (ERK1/2), and c-Jun N-terminal kinase 1/2 (JNK1/2) were determined by Western blotting (n=4). The same samples of rats in control group were collected for related detection at the same time point as above. Data were processed with one-way analysis of variance and SNK test. Results: (1) The serum content of FITC-dextran of rats in pure sepsis group was significantly higher than that in control group (P<0.01), while the serum content of FITC-dextran of rats in NHE1 inhibitor group was significantly lower than that in pure sepsis group (P<0.01). Compared with that in control group, infiltration of a large number of inflammatory cells, ulcer and necrosis of intestinal mucosa of rats in pure sepsis group were observed. The injury condition of intestine of rats in NHE1 inhibitor group was better than that in pure sepsis group. (2) The serum content of IL-6, TNF-α, and MPO of rats in pure sepsis group was (387±42) and (164.7±10.1) ng/mL, and (7.5±1.5) U/mL, respectively, significantly higher than that in control group [(75±17) and (13.1±6.5) ng/mL, and (2.3±0.7) U/mL, respectively, with P values below 0.01]. The serum content of IL-6, TNF-α, and MPO of rats in NHE1 inhibitor group was (176±37) and (64.9±9.3) ng/mL, and (5.9±0.8) U/mL, respectively, which was significantly lower than that in pure sepsis group (with P values below 0.01). (3) The content of IL-6, TNF-α, and MPO in intestinal tissue of rats in pure sepsis group was (190±13) and (172.8±29.7) ng/mL, and (8.7±1.5) U/mL, respectively, significantly higher than that in control group [respectively (20±3) and (11.9±2.3) ng/mL, and (2.9±0.3) U/mL, with P values below 0.01]. The content of IL-6, TNF-α, and MPO of intestinal tissue of rats in NHE1 inhibitor group was (35±6) and (45.2±6.1) ng/mL, and (5.3±0.6) U/mL, respectively, significantly lower than that in pure sepsis group (with P values below 0.01). (4) The protein expression of NF-κB-p65 and phosphorylation levels of p38MAPK and ERK1/2 in intestinal tissue of rats in pure sepsis group were significantly higher than those in control group (with P values below 0.01); the protein expression of NF-κB-p65 and the phosphorylation level of p38MAPK in intestinal tissue of rats in NHE1 inhibitor group were significantly lower than those in pure sepsis group (with P values below 0.01); phosphorylation levels of JNK1/2 in intestinal tissue of rats in the three groups were similar (with P values above 0.05). Conclusions The inhibition of NHE1 can significantly alleviate the intestinal injury, and the mechanisms may be attributed to the regulation of NF-κB and p38MAPK signal pathway, resulting in inhibition of the inflammatory response of intestinal tract.

Objective: To investigate the interventional effects of BAY11-7082 on lung inflammatory response at the early stage and acute lung injury of rats with severe burns. Methods: (1) Experiment 1. Twelve Sprague-Dawley (SD) rats were divided into control (C) group and burn (B) group according to the random number table, with 3 rats in group C and 9 rats in group B. Rats in group C did not receive any special treatment. Rats in group B were inflicted with 30% total body surface area full-thickness burn on the back. Immediately after injury, rats in group B were intraperitoneally injected with normal saline in the dosage of 50 mL/kg. Abdominal aorta blood and lung tissue samples were collected from three rats in group B at post injury hour (PIH) 12, 24, and 48, respectively. The interleukin-1β (IL-1β) and the IL-18 content of serum were determined with enzyme-linked immunosorbent assay. The mRNA expressions of IL-1β and IL-18 in lung tissue were determined with real-time fluorescent quantitative reverse transcription polymerase chain reaction (RT-PCR). Sample collection and determination in rats of group C were performed as above. (2) Experiment 2. Eighteen SD rats were divided into control (C) group, simple burn (SB) group, and BAY11-7082 intervention (BI) group according to the random number table, with 6 rats in each group. Rats in group C did not receive any special treatment. Rats in groups SB and BI were inflicted with injury as in experiment 1. Immediately after injury, rats in group SB were intraperitoneally injected with normal saline in the dosage of 50 mL/kg, and those in group BI with 8 mg/mL (final mass concentration) BAY11-7082 solution in the dosage of 50 mL/kg. Lung tissue and bronchoalveolar lavage fluid (BALF) of rats with burns were collected at the optimal observation time point concluded from experiment 1. The morphology of lung tissue was observed with hematoxylin-eosin staining, and the pathological damage of lung tissue was graded. The myeloperoxidase (MPO) content of lung tissue and the total protein content of BALF were detected by microplate reader. The protein expressions of nucleotide-binding oligomerization domain-like receptor-3 (NLRP3) and cysteine-aspartic proteases 1 (caspase-1) in lung tissue were determined with Western-blotting. The mRNA expressions of IL-1β, IL-18, NLRP3, and caspase-1 in lung tissue were determined with real-time fluorescence quantitative RT-PCR. Sample collection and determination in rats of group C were performed as above. Data were processed with one-way analysis of variance and LSD-t test. Results: (1) The IL-1β and IL-18 content of serum in rats of group B at PIH 12, 24, and 48 were significantly higher than those of group C (t=10.55, 22.05, 12.47, 10.60, 15.22, 11.94, P<0.01). The mRNA expressions of IL-1β and IL-18 in rats of group B at PIH 12, 24, and 48 were significantly higher than those of group C (t=3.62, 7.19, 5.28, 3.20, 12.62, 7.31, P<0.05 or P<0.01). PIH 24 was the optimal observation time point for the following experiment. (2) At PIH 24, compared with those in group SB, the inflammatory cell infiltration and erythrocyte exudates of alveolar in group BI were obviously reduced, and the pulmonary interstitial edema obviously subsided. The pathological damage score of lung tissue in rats of group SB was (9.00±1.00) points, significantly higher than (1.10±0.26) points of group C (t=13.23, P<0.01). The pathological damage score of lung tissue in rats of group BI was (4.93±0.70) points, which was significantly lower than that of group SB (t=5.76, P<0.01) but still significantly higher than that of group C (t=8.84, P<0.01). At PIH 24, the MPO content of lung tissue and the total protein content of BALF in rats of group SB were (1.83±0.15) U/mg and (1.39±0.20) mg/mL, respectively, significantly higher than (0.51±0.10) U/mg and (0.44±0.05) mg/mL of group C (t=12.50, 7.86, P<0.01). The MPO content of lung tissue and the total protein content of BALF in rats of group BI were (0.91±0.12) U/mg and (0.60±0.10) mg/mL, respectively, significantly lower than those of group SB (t=8.36, 6.06, P<0.01). At PIH 24, the protein expressions of NLRP3 and caspase-1 in lung tissue of rats of group SB were 3.10±0.09 and 2.99±0.30, respectively, significantly higher than 1.00 and 1.00 of group C (t=9.06, 11.28, P<0.01). The protein expressions of NLRP3 and caspase-1 in lung tissue of rats of group BI were 1.13±0.08 and 1.81±0.11, respectively, significantly lower than those of group SB (t=7.24, 3.91, P<0.05 or P<0.01). At PIH 24, the mRNA expressions of IL-1β, IL-18, NLRP3, and caspase-1 in lung tissue of rats in group SB were 5.0±0.4, 3.32±0.21, 3.54±0.42, and 6.3±1.0, respectively, significantly higher than 1.0, 1.00, 1.00, and 1.0 of group C (t=13.97, 14.14, 11.78, 7.13, P<0.01). The mRNA expressions of IL-1β, IL-18, NLRP3, and caspase-1 in lung tissue of rats in group BI were 2.6±0.5, 2.00±0.28, 1.39±0.21, and 2.5±0.5, respectively, significantly lower than those of group SB (t=7.11, 5.80, 9.99, 4.65, P<0.05 or P<0.01). Conclusions Applying BAY11-7082 at the early stage of acute lung injury of rats with severe burn can reduce the expression of caspase-1, decrease the levels of IL-1β and IL-18, and decrease the MPO content of lung tissue and the total protein content of BALF through inhibiting NLRP3, thus alleviating the lung inflammatory response and lung injury.

Objective: To observe the effects of the method of combining free superficial temporal fascia flaps with skin grafts in repairing deep wounds in posterior ankle region of extensively burned patients. Methods: From September 2013 to February 2017, 11 extensively burned patients with deep tissue defects in posterior ankle region were treated in our unit. Two patients had tissue defects in bilateral posterior ankle regions. The wound sizes ranged from 5.8 cm×4.6 cm to 11.7 cm×5.2 cm. Free superficial temporal fascia flaps with the same sizes as the wounds were designed and resected to repair wounds in posterior ankle regions after debridement. The proximal end of superficial temporal veins and posterior tibial veins were performed with end-to-end anastomosis, and superficial temporal arteries and posterior tibial arteries were performed with end-to-side anastomosis. Skin grafts were resected to cover the superficial temporal fascia flaps according to patients′ condition of donor sites. The donor sites were sutured directly. Results: All fascial flaps in 11 patients survived, while 2 skin grafts had partial necrosis, and they healed after secondary skin graft. All patients were followed up for 6 to 13 months, and the shape and function of the operation sites in all patients recovered well. Conclusions The method of combining free superficial temporal fascia flaps with skin grafts can repair deep wounds in posterior ankle regions of extensively burned patients. It has the advantages of good appearances in the recipient sites, strong resistances to infection of fascia flaps, minimal damages to the donor sites, short course of disease, and good prognosis of patients.

Objective: To explore the choice of the donor site of flap and the repair method of secondary wound of flap donor site in tissue repair and reconstruction operation. Methods: From January 2014 to September 2018, 62 cases of scar contracture deformity, 15 cases of skin tumor, 20 cases of skin and soft tissue injury, and 25 cases of chronic wound were admitted to the Burn Center of People′s Liberation Army of First Affiliated Hospital of Air Force Medical University, with 84 males and 38 females, aged from 3 to 89 years. Four repair strategies adopted for tissue repair and reconstruction and good repair of the donor site of flap were as follows: designing the flap rationally according to the condition around the wound or the size and shape of wound, choosing pre-expanded technique of the donor site of flap for repair of scar deformity optimally, making full use of the surrounding condition of flap donor site, and repaired with the distal flap, i. e. replacing the important site with secondary site. The donor site of flap was repaired by direct suture or peripheral flap and distal flap. The wound size of patients ranged from 3.0 cm×2.0 cm to 20.0 cm×18.0 cm, and the flap area ranged from 3.5 cm×2.0 cm to 25.0 cm×22.0 cm. The survival condition of flap, healing condition of donor site and recipient site, and follow-up condition of donor site and recipient site were recorded. Results: Wounds of 122 patients were repaired with a total of 148 flaps designed by the above four repair strategies. All the flaps survived well, and the wound and flap donor site healed well. Follow-up for 3 to 36 months showed that the shape and function of recipient site and flap donor site were satisfactory. Conclusions According to the specific condition of the wound and anatomical structure of the surrounding tissue of flap donor site, overall surgical design with flexibility and personalization can achieve effects of good repair of the wound and reduce the secondary damage of flap donor site.

If the abdominal pedicled flaps are not well fixed after repair of deep electric burn wounds in hands, many problems such as poor blood supply may occur. In order to solve the above problems, we designed and manufactured the individualized low temperature thermoplastic plate combined with special abdominal band to fix abdominal pedicled flaps for repairing of 17 patients (12 males and 5 females, aged 2-35 years) with deep electric burn wounds in hands from February 2016 to August 2018, and achieved the desired results. The shoulder joint, elbow joint, and wrist joint were fixed by low temperature thermoplastic plate according to the 1/2 circumference of the patient′s side chest and upper arm, and the braking of wrist joint and elbow joint was strengthened by special abdominal band. Application of the combined method of fixing abdominal pedicled flaps in repairing deep electric burn wounds in hands has high success rate of flap transplantation. It is simple to make and practical, and worthy of clinical promotion.

Objective: Strategy for wound repair of skin and soft tissue defect and systematic rehabilitation treatment for functional reconstruction of patients with severe burn or trauma on knees. Methods: From January 2015 to October 2016, 26 patients with skin and soft tissue defect on knees after severe burn or trauma were hospitalized in our unit. Among these patients, 14 patients had patellar ligament defect, and 16 patients had knee joint capsule defect. Wound debridement was operated on 1 to 3 days after admission. After debridement, the area of skin and soft tissue defect ranged from 10 cm×7 cm to 42 cm×18 cm. Vacuum sealing drainage (VSD) treatment was performed after debridement, and flap transplantation operation was performed after VSD treatment for 5 to 7 days. Defects of nine patients were treated with local rotation flaps. Seven patients with skin and soft tissue defects on knees and knee joint capsule defects of 5 cm×3 cm to 9 cm×7 cm were treated with free anterolateral femoral flaps combined with fascia lata. Ten patients with skin and soft tissue defects on knees and patellar ligament defects of 6 cm×3 cm to 12 cm×4 cm were treated with free anterolateral femoral flaps combined with iliotibial tract. The area of flaps ranged from 11 cm×9 cm to 22 cm×15 cm. After flap transplantation operation, functional reconstruction of knee joint was carried out according to early, continuous, and sequential systematic rehabilitation treatment strategy. The pain degree and function of knee joint of patients were scored by the International Knee Documentation Committee (IKDC) Knee Evaluation Form before operation and 12 months after operation. The knee joint flexion and extension degrees of patients were measured by joint protractor in 2 weeks and 12 months after operation. The color Doppler ultrasound was used to evaluate integrity of knee joint capsule and continuity of patellar ligament of patients in 6 and 12 months after operation. Results: All flaps of 26 patients survived well, and wounds healed completely after the operation. Distal parts of flaps of 2 patients treated with free anterolateral femoral flaps had local necrosis after the operation, and their wounds healed after debridement and transplantation of autologous intermediate split-thickness skin graft of thigh. The IKDC Knee Evaluation Form score of patients was (79±8) points in 12 months after operation, which was significantly higher than (64±7) points before operation (t=7.20, P<0.05). The flexion degree of knee joint of patients was (117±10)° in 12 months after operation, which was significantly larger than (35±8)° in 2 weeks after operation (t=32.65, P<0.05). The extension degree of knee joint of patients was (12±9)° in 12 months after operation, which was significantly smaller than (61±9)° in 2 weeks after operation (t=19.63, P<0.05). In 6 and 12 months after operation, 9 patients treated with local rotation flaps had good integrity of knee joint capsule and continuity of patellar ligament; 7 patients treated with free anterolateral femoral flaps and fascia lata had good integrity of knee joint capsule; 10 patients treated with free anterolateral femoral flaps and iliotibial tract had good continuity of patellar ligament. During follow-up of 12 months, all flaps survived well; knees of all patients had good appearance; knee joints functioned normally. Conclusions Good appearance and function of knees can be achieved by repairing wound of skin and soft tissue defect on knees after severe burn or trauma with local rotation flaps or free anterolateral femoral flaps with fascia lata or iliotibial tract plus systematic rehabilitation treatment of knee joint in early stage after flap transplantation operation.

Objective:To explore the effect of free transplantation of composite tissue flap from the anterior lateral aspect of the femur in repairing head skin defects with artificial dural exposure infection.Methods:A retrospective study was conducted on 13 patients admitted to the First Affiliated Hospital of Air Force Military Medical University from April 2018 to August 2020 with craniotomy complications, including craniotomy skin and soft tissue defects combined with artificial dural exposure and infection. After preoperative anti infection treatment, the neurosurgery department participated in debridement and removed the artificial dura mater as much as possible during the operation. A composite tissue flap carrying the fascia lata was designed for the anterior lateral aspect of the thigh, and the flap artery and vein were anastomosed with the superficial temporal artery and superficial temporal vein/middle temporal vein respectively. The defect of the dura mater was repaired with the fascia lata with blood supply. The flap was used to seal the wound, and the donor site was directly sutured or transplanted with autologous medium thick skin graft. The postoperative blood supply and survival of the flap, the presence of cerebrospinal fluid leakage, and the healing of the donor site were observed; The observation of dural integrity and postoperative effects of skull reconstruction using cranial magnetic resonance imaging was followed up.Results:Among the 13 patients in this group, 11 patients had their artificial dura mater completely removed, while 2 patients were not completely removed due to severe adhesion. Among them, 1 patient had a residual area of 0.8 cm×1 cm, and the other had 3 residual areas, with a maximum area of 0.5 cm×0.7 cm; All transplanted skin flaps survived, with 12 cases achieving primary healing and 1 case of partial wound rupture after suture removal, which healed after conservative dressing change; All patients had no cerebrospinal fluid leakage; There was one case of partial necrosis of the graft in the donor site, which healed after supplementing the graft; Thirteen patients underwent cranial magnetic resonance imaging at 3-6 months postoperatively, all of which showed intact dura mater; Among them, 8 patients have completed skull reconstruction surgery, and all of them have healed well after reconstruction, with a good appearance of the surgical area.Conclusions:For wounds with head skin defects and exposed artificial dura mater infection, free transplantation of the anterior lateral composite tissue flap carrying the fascia lata can effectively cover the wound and repair the dura mater defect, achieve good function and appearance, and create favorable conditions for later skull reconstruction.