2019 Vol. 35, No. 2
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2019, 35(2): 81-85.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.001
Abstract
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Abstract:
Objective Burn rehabilitation medicine in China has made great progress in recent years. This article reviews the development history of burn rehabilitation medicine in China from three stages of the initial period, the starting stage, and the development period by looking back to the history. Besides, the article is written according to the milestone events in the development of burn rehabilitation medicine, such as article publishing, guideline making, monograph publication, establishment of rehabilitation association, rehabilitation conference holding. By summarizing successful experience in the past and analyzing the challenge we face, the authors wish all the colleagues committed to burn prevention and treatment work together to make the burn rehabilitation medicine in China better in the future.
Objective Burn rehabilitation medicine in China has made great progress in recent years. This article reviews the development history of burn rehabilitation medicine in China from three stages of the initial period, the starting stage, and the development period by looking back to the history. Besides, the article is written according to the milestone events in the development of burn rehabilitation medicine, such as article publishing, guideline making, monograph publication, establishment of rehabilitation association, rehabilitation conference holding. By summarizing successful experience in the past and analyzing the challenge we face, the authors wish all the colleagues committed to burn prevention and treatment work together to make the burn rehabilitation medicine in China better in the future.
2019, 35(2): 86-89.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.002
Abstract:
Along with the development of society and the change of disease spectrum, chronic wound is gradually becoming the core of burn and plastic surgery field. Although there have been some progresses in the diagnosis and treatment technology, the management strategy of chronic wound is still in the traditional mode stage. The development of internet of things, cloud computing, big data, artificial intelligence, and other emerging technologies is changing with each passing day, and they have rapidly penetrated into the health care field. To explore the application prospect of emerging technology in the diagnosis and treatment management of chronic wound and to plan its strategy and mode in the diagnosis and treatment of chronic wound can further promote development of discipline of burns.
Along with the development of society and the change of disease spectrum, chronic wound is gradually becoming the core of burn and plastic surgery field. Although there have been some progresses in the diagnosis and treatment technology, the management strategy of chronic wound is still in the traditional mode stage. The development of internet of things, cloud computing, big data, artificial intelligence, and other emerging technologies is changing with each passing day, and they have rapidly penetrated into the health care field. To explore the application prospect of emerging technology in the diagnosis and treatment management of chronic wound and to plan its strategy and mode in the diagnosis and treatment of chronic wound can further promote development of discipline of burns.
2019, 35(2): 90-94.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.003
Abstract:
Objective To explore the characteristics of patients with tuberculous wounds and non-tuberculous chronic refractory wounds in single center. Methods From January 2010 to June 2017, 43 patients with tuberculous wounds and 44 patients with non-tuberculous chronic refractory wounds admitted to the Department of Burns and Plastic Surgery of the Eighth Medical Center of the General Hospital of the Chinese People′s Liberation Army were conforming to the inclusion criteria. The patients were assigned to tuberculous wound group and non-tuberculous wound group, respectively, and their clinical records were retrospectively analyzed. The gender, place of residence, history of trauma, time of wound formation, time of wound diagnosis, number and length of hospital stay, age, wound site, wound area, sinus occurrence, number of dressing change, number of operation, vacuum sealing drainage (VSD) treatment, recovery, source of medical expense, expense paid by social basic medical insurance and the self-payment of patients in the 2 groups were investigated. Data were processed with independent samplet test and chi-square test.
Results (1) Except for gender (χ 2=0.019, P >0.05), there were significantly statistical differences in place of residence, history of trauma, time of wound formation, time of wound diagnosis, number and length of hospital stay between patients in tuberculous wound group and non-tuberculous wound group (χ 2=4.535, 27.651, t =7.252, 16.131, 4.663, 7.416, P <0.05 or P <0.01). (2) There was no statistically significant difference in the composition ratio of age between patients in tuberculous wound group and non-tuberculous wound group (χ 2=11.522, P >0.05). (3) The wounds of patients in tuberculous wound group were more common in the chest, and the wounds of patients in non-tuberculous wound group were more common in the lower limbs. There was statistically significant difference in the composition ratio of the wound sites between patients in the two groups (χ 2=28.450, P <0.01). (4) There were statistically significant differences in wound area, sinus occurrence, number of dressing change, number of operation between patients in tuberculous wound group and non-tuberculous wound group (t =-8.524, 9.846, -15.426, 4.663, P <0.01). There were no statistically significant differences in VSD treatment and recovery between patients in the two groups (χ 2=0.032, 0.111, P >0.05). (5) The medical expenses of patients in tuberculous wound group from social basic medical insurance, free medical service, the self-paid, and military medical services accounted for 48.8% (21/43), 7.0% (3/43), 39.5% (17/43), and 4.7% (2/43), respectively. The medical expenses of patients in non-tuberculous wound group from social basic medical insurance, free medical service, the self-paid, and military medical services accounted for 59.1% (26/44), 4.5% (2/44), 29.5% (13/44), and 6.8% (3/44), respectively. There was no statistically significant difference in the composition ratio of sources of medical expense between patients in the two groups (χ 2=1.154, P >0.05). (6) There were statistically significant differences in expenses for diagnosis, medicine, surgery, examination, laboratory test, and bed, and total expenses paid by social basic medical insurance and the self-payment between patients in tuberculous wound group and non-tuberculous wound group (t =45.051, 39.995, 64.212, 32.584, 8.754, 43.991, 15.671, 17.640, 65.155, 35.546, 35.903, -4.329, 3.344, 12.984, P <0.01).
Conclusions Compared with those of patients with non-tuberculous chronic refractory wounds, the tuberculous wounds of patients have longer formation time, the diagnosis and treatment of the wounds are difficult, their wounds are mostly distributed in the chest and often accompanied by sinus formation, and patients with the wounds have long hospital stay and high medical expenses. Besides, the medical expenses for treating wounds of patients in the two groups are mainly paid by social basic medical insurance and the patients themselves.
Objective To explore the characteristics of patients with tuberculous wounds and non-tuberculous chronic refractory wounds in single center. Methods From January 2010 to June 2017, 43 patients with tuberculous wounds and 44 patients with non-tuberculous chronic refractory wounds admitted to the Department of Burns and Plastic Surgery of the Eighth Medical Center of the General Hospital of the Chinese People′s Liberation Army were conforming to the inclusion criteria. The patients were assigned to tuberculous wound group and non-tuberculous wound group, respectively, and their clinical records were retrospectively analyzed. The gender, place of residence, history of trauma, time of wound formation, time of wound diagnosis, number and length of hospital stay, age, wound site, wound area, sinus occurrence, number of dressing change, number of operation, vacuum sealing drainage (VSD) treatment, recovery, source of medical expense, expense paid by social basic medical insurance and the self-payment of patients in the 2 groups were investigated. Data were processed with independent sample
2019, 35(2): 95-103.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.004
Abstract:
Objective To investigate the effects of basic fibroblast growth factor (bFGF) on healing ofMycobacterium tuberculosis infective wound in New Zealand rabbit after debridement.
Methods Thirty-two New Zealand rabbits (3 to 4 months old, no matter male or female) were intradermally injected with 0.1 mL of complete Freund′s adjuvant on the buttocks. Six weeks later, each rabbit was injected with 0.5 mL 5×107 colony forming unit/mL Bacillus Calmette -Guerin on both sides of the back to reproduce the model of Mycobacterium tuberculosis infective wound in New Zealand rabbit. After successful modeling, the 32 rabbits were divided into growth factor (GF) group, antituberculosis drug (AD) group, combined treatment (CT) group, and blank control (BC) group according to the random number table, with 8 rabbits in each group. After a complete debridement, the wounds of rabbits in group GF were treated with recombinant bovine bFGF gel (300 IU/cm2, about 0.45 g for each wound), the wounds of rabbits in group AD were covered with gauze which was impregnated with 6 mL isoniazid injection and 0.15 g rifampicin powder-injection, the wounds of rabbits in group CT were covered with gauze which was impregnated with isoniazid injection and rifampicin powder-injection after being treated with recombinant bovine bFGF gel as before, the wounds of rabbits in group BC were covered with sterile gauze, with dressing change of once every two days until the wounds were completely healed. Immediately after surgery and on post surgery day (PSD) 7, 14, 21, and 28, the wounds of rabbits in each group were observed with naked eyes and photos. On PSD 7, 14, 21, and 28, the wound healing rate was calculated and the complete healing time of wound was recorded. On PSD 7, 14, 21, and 28, the tissue samples of wound edge were collected for histomorphological observation with hematoxylin and eosin staining and Masson staining. On PSD 21, the number of microvessels was counted with immunohistochemical method. On PSD 7, 14, 21, and 28, the content of hydroxyproline in wound edge was determined by enzyme-linked immunosorbent assay. The numbers of samples of above-mentioned experiments were all 8. Data were processed with analysis of variance for repeated measurement, analysis of variance of factorial design, one-way analysis of variance, least significant difference test and Bonferroni correction.
Results (1) The rabbits in four groups all survived to the end of experiment. Immediately after surgery, edema was observed in basal wounds of rabbits in the four groups. On PSD 7, the wounds of rabbits in the 4 groups were contracted with scabs and less edema. The wounds of rabbits in groups GF and CT became redder. On PSD 14, the wounds of rabbits in the 4 groups contracted obviously. There were no obvious exudates in wounds of rabbits in groups AD and CT, while 1 wound of rabbit in group GF and 2 wounds of rabbits in group BC became red and swelling with purulent exudates. On PSD 21, wounds of rabbits in groups GF and CT were basically healed, while 2 wounds of rabbits in group BC healed slowly with purulent secretion. On PSD 28, wounds of rabbits in the 4 groups were basically healed, while 2 wounds of rabbits in group BC hardly healed with redness and swelling. (2) From PSD 7 to 28, the wound healing rates of rabbits in groups GF, AD, and CT were significantly higher than those in group BC (P <0.05). On PSD 14 and 21, the wound healing rates of rabbits in groups GF and CT were significantly higher than those in group AD (P <0.05). From PSD 7 to 28, the wound healing rates of rabbits in group GF were close to those in group CT (P >0.05). (3) The complete healing time of wounds of rabbits in groups GF, AD, and CT was significantly shorter than that in group BC (P <0.05). The complete healing time of wounds of rabbits in groups GF and CT was significantly shorter than that in group AD (P <0.05). The complete healing time of wounds of rabbits in group GF was close to that in group CT (P >0.05). (4) On PSD 7, a large number of inflammatory cells infiltration were observed in wound tissue of rabbits in the 4 groups and a few epithelial cells were observed in wound tissue of rabbits in groups GF, AD, and CT. On PSD 14, more epithelial cells were observed in wound tissue of rabbits in groups GF and CT, and an obvious reduction of inflammatory cells infiltration was observed in wound tissue of rabbits in groups AD and CT. On PSD 21, there was a complete wound tissue structure and distinctive nuance of dyeing in wound tissue of rabbits in groups GF and CT while thinner new epithelium in wound tissue of rabbits in groups AD and BC, and inflammatory cell infiltration was observed in wound tissue of rabbits in group BC. On PSD 28, there was a complete wound tissue structure in wound tissue of rabbits in the 4 groups, the new epithelium in wound tissue of rabbits in groups GF, AD, and CT was thicker than that in group BC. (5) On PSD 7 and 14, the quantity of collagen fibers in wound tissue of rabbits in groups GF and CT was larger than that in the other two groups. On PSD 21, a large quantity of fibroblasts and well reorganized collagen fibers were observed in wound tissue of rabbits in groups GF and CT, a moderate quantity of fibroblasts and collagen fibers in a random arrangement were observed in wound tissue of rabbits in group AD, and a little quantity of fibroblasts and collagen fibers were observed in wound tissue of rabbits in group BC. On PSD 28, the quantity of collagen fibers in wound tissue of rabbits in the 4 groups was close to that of normal skin tissue, and the collagen fibers performed more well reorganized in wound tissue of rabbits in groups GF and CT. (6) On PSD 21, the numbers of microvessels per 200-time visual field in wound edge of rabbits in groups GF (31.6±1.2), AD (27.5±1.3), and CT (32.8±1.6) were significantly higher than the number in group BC (22.3±1.7, P <0.05). The numbers of microvessels in wound edge of rabbits in groups GF and CT were significantly higher than the number in group AD (P <0.05). The number of microvessels in wound edge of rabbits in group GF was close to that in group CT (P >0.05). (7) On PSD 7 and 28, there were no statistically significant differences in content of hydroxyproline in wound edge of rabbits in the 4 groups (F =0.916, 1.752, P >0.05). On PSD 14 and 21, the content of hydroxyproline in wound edge of rabbits in groups GF, AD, and CT was significantly higher than that in group BC (P <0.05). The content of hydroxyproline in the wound edge of rabbits in groups GF and CT was significantly higher than that in group AD (P <0.05). The content of hydroxyproline in the wound edge of rabbits in group GF was close to that in group CT (P >0.05).
Conclusions bFGF can be used solely or combined with AD to promote Mycobacterium tuberculosis infective wound healing in New Zealand rabbit after complete debridement of wound, which is better than single use of AD.
Objective To investigate the effects of basic fibroblast growth factor (bFGF) on healing of
2019, 35(2): 104-109.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.005
Abstract:
Objective To explore the regulatory mechanism of E2F1 transcription factor on M2 macrophages in full-thickness skin defect wounds of mice. Methods E2F1 gene knockout heterozygotes C57BL/6 mice and wild-type C57BL/6 mice were introduced and self-reproduced. Two weeks after birth, E2F1 gene knockout homozygotes mice and wild-type mice were identified by polymerase chain reaction (PCR). Twelve identified 6-8 weeks old male E2F1 gene knockout homozygotes C57BL/6 mice and wild-type C57BL/6 mice were selected respectively according to the random number table and set as E2F1 gene knockout group and wild-type group. A full-thickness skin defect wound was made on the back of each mouse. On post injury day (PID) 2 and 7, 6 mice in each group were selected according to the random number table and sacrificed, and the wound tissue was excised. The expression of CD68 and CD206 double positive M2 macrophages was observed by immunofluorescence method, and the percentage of CD206 positive cells was calculated. The protein expression of CD206 was detected by Western blotting. The mRNA expression of arginase 1 was detected by real-time fluorescent quantitative reverse transcription PCR (RT-PCR). Wound tissue specimens of the two groups on PID 7 were obtained, and the protein and mRNA expressions of peroxisome proliferator-activated receptor gamma (PPAR-γ) were detected by Western blotting and real-time fluorescent quantitative RT-PCR respectively. The above-mentioned experiments were repeated four times. Three specimens of wound tissue of mice in wild-type group on PID 7 were obtained to detect the relationship between E2F1 and PPAR-γ by co-immunoprecipitation and Western blotting, and this experiment was repeated two times. Data were processed with unpairedt test.
Results The size of PCR products of E2F1 gene knockout homozygotes C57BL/6 mice and wild-type C57BL/6 mice were 227 and 172 bp respectively, which were the same as those of the designed DNA fragments. On PID 2 and 7, the number of CD68 and CD206 double positive M2 macrophages in the wound tissue of mice in E2F1 gene knockout group was more than that of wild-type group, and the percentages of CD206 positive cells in the wound tissue of mice in E2F1 gene knockout group were (0.234±0.032)% and (0.584±0.023)% respectively, which were significantly higher than (0.129±0.017)% and (0.282±0.071)% of wild-type group (t =3.29, 3.54, P <0.05). On PID 2 and 7, the protein expression of CD206 in the wound tissue of mice in E2F1 gene knockout group were 1.00±0.23 and 1.63±0.26 respectively, which were significantly higher than 0.43±0.06 and 0.97±0.08 of wild-type group (t =2.41, 2.45, P <0.05). On PID 2 and 7, the mRNA expressions of arginase 1 in the wound tissue of mice in E2F1 gene knockout group were 0.482±0.105 and 0.195±0.031 respectively, which were significantly higher than 0.163±0.026 and 0.108±0.017 of wild-type group (t =3.04, 2.86, P <0.05). On PID 7, the protein and mRNA expressions of PPAR-γ in the wound tissue of mice in E2F1 gene knockout group were 0.61±0.12 and 0.51±0.13 respectively, which were significantly higher than 0.20±0.04 and 0.20±0.04 of wild-type group (t =3.36, 2.86, P <0.05). On PID 7, detection of the wound tissue of mice in wild-type group showed that PPAR-γ had unidirectional effect on E2F1.
Conclusions E2F1 transcription factor affects the polarization of M2 macrophages by inhibiting the expression of PPAR-γ, thereby inhibiting the healing process of full-thickness skin defect wounds in mice.
Objective To explore the regulatory mechanism of E2F1 transcription factor on M2 macrophages in full-thickness skin defect wounds of mice. Methods E2F1 gene knockout heterozygotes C57BL/6 mice and wild-type C57BL/6 mice were introduced and self-reproduced. Two weeks after birth, E2F1 gene knockout homozygotes mice and wild-type mice were identified by polymerase chain reaction (PCR). Twelve identified 6-8 weeks old male E2F1 gene knockout homozygotes C57BL/6 mice and wild-type C57BL/6 mice were selected respectively according to the random number table and set as E2F1 gene knockout group and wild-type group. A full-thickness skin defect wound was made on the back of each mouse. On post injury day (PID) 2 and 7, 6 mice in each group were selected according to the random number table and sacrificed, and the wound tissue was excised. The expression of CD68 and CD206 double positive M2 macrophages was observed by immunofluorescence method, and the percentage of CD206 positive cells was calculated. The protein expression of CD206 was detected by Western blotting. The mRNA expression of arginase 1 was detected by real-time fluorescent quantitative reverse transcription PCR (RT-PCR). Wound tissue specimens of the two groups on PID 7 were obtained, and the protein and mRNA expressions of peroxisome proliferator-activated receptor gamma (PPAR-γ) were detected by Western blotting and real-time fluorescent quantitative RT-PCR respectively. The above-mentioned experiments were repeated four times. Three specimens of wound tissue of mice in wild-type group on PID 7 were obtained to detect the relationship between E2F1 and PPAR-γ by co-immunoprecipitation and Western blotting, and this experiment was repeated two times. Data were processed with unpaired
Chen Bin,
Kuang Fang,
Li Xiaojian,
Zhang Zhi,
Deng Zhongyuan,
Zhang Xuhui,
Zhang Tao,
Zhong Xiaomin,
Tang Wenbin,
Liu Changling
2019, 35(2): 110-115.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.006
Abstract:
Objective To analyze the causes of complication of early acute kidney injury (AKI) in four severely burned patients, and to explore the related treatment methods. Methods The clinical data of 4 patients with severe burn complicated with early AKI admitted to Guangzhou Red Cross Hospital Affiliated to Medical College of Jinan University (hereinafter referred to as our hospital) from June 2014 to December 2017 were retrospectively analyzed. All the patients were male, aged 23-33 (30±5) years old, with depth of burns ranged from deep partial-thickness to full-thickness, complicated with myofascial compartment syndrome of extremities and varying degrees of striated muscle injury, and treated in other hospitals before transfer to our hospital. The patients were numbered from small to large according to the total burn area. The total burn area of patients No. 1, 2, 3, and 4 was 10%, 80%, 90%, and 95% total body surface area respectively, their occurrence time of early AKI was 48, 11, 29, and 48 hours after injury respectively, and their time of arriving our hospital was 60, 11, 29, and 144 hours after injury respectively. Hypovolemic shock occurred in patients No. 2 and 3 at admission to our hospital. All the patients received continuous renal replacement therapy (CRRT) after admission to our hospital. Under the support of hemodynamic monitoring and organ function monitoring, the limbs complicated with myofascial compartment syndrome were incised, thorough decompression exploration was performed, and necrotic muscle tissue was removed or amputation was performed. After escharectomy and decompression of limbs, fresh granulation wounds were formed by temporarily covering wounds with Jieya dressing skin or pig skin, multiple debridements, and vacuum sealing drainage. Fresh granulation wounds and other wounds underwent staged eschar excision and shaving were covered with autologous Meek skin graft, particulate skin graft, reticular skin graft and small skin graft respectively. The treatment outcome, CRRT time, operation times, time of recovery of serum creatinine and myoglobin, length of hospital stay, and follow-up were recorded. Results All the 4 patients were cured after transfer to our hospital. Among them, totally 5 limbs of patients No. 1 and No. 4 underwent amputation because of complication of myofascial compartment syndrome and a large amount of necrotic muscle which could not be preserved. Patients No. 1, 2, 3, and 4 were treated with CRRT for 19, 35, 14, and 25 days respectively and performed with operation for 5, 6, 10, 8 times respectively. Serum creatinine of patients No. 1, 2, 3, and 4 returned to normal on 22, 35, 37, and 48 days after transfer respectively, and their serum myoglobin returned to normal on 18, 28, 25, and 30 days after transfer respectively. Patients No. 1, 2, 3, and 4 were hospitalized for 52, 105, 148, and 156 days and discharged after basic wound healing. Follow-up for 1 to 36 months showed no abnormal renal function in 4 patients. Conclusions The early AKI in patients No. 1 and 4 was caused by rhabdomyolysis after severe burn complicated with myofascial compartment syndrome, while that of the other 2 cases were also related to hypovolemic shock and poor renal perfusion. The success rate of early AKI treatment in severely burned patients can be effectively improved by removing the causes of diseases at the same time of CRRT and actively treating burn wounds under the support of organ function and hemodynamic monitoring.
Objective To analyze the causes of complication of early acute kidney injury (AKI) in four severely burned patients, and to explore the related treatment methods. Methods The clinical data of 4 patients with severe burn complicated with early AKI admitted to Guangzhou Red Cross Hospital Affiliated to Medical College of Jinan University (hereinafter referred to as our hospital) from June 2014 to December 2017 were retrospectively analyzed. All the patients were male, aged 23-33 (30±5) years old, with depth of burns ranged from deep partial-thickness to full-thickness, complicated with myofascial compartment syndrome of extremities and varying degrees of striated muscle injury, and treated in other hospitals before transfer to our hospital. The patients were numbered from small to large according to the total burn area. The total burn area of patients No. 1, 2, 3, and 4 was 10%, 80%, 90%, and 95% total body surface area respectively, their occurrence time of early AKI was 48, 11, 29, and 48 hours after injury respectively, and their time of arriving our hospital was 60, 11, 29, and 144 hours after injury respectively. Hypovolemic shock occurred in patients No. 2 and 3 at admission to our hospital. All the patients received continuous renal replacement therapy (CRRT) after admission to our hospital. Under the support of hemodynamic monitoring and organ function monitoring, the limbs complicated with myofascial compartment syndrome were incised, thorough decompression exploration was performed, and necrotic muscle tissue was removed or amputation was performed. After escharectomy and decompression of limbs, fresh granulation wounds were formed by temporarily covering wounds with Jieya dressing skin or pig skin, multiple debridements, and vacuum sealing drainage. Fresh granulation wounds and other wounds underwent staged eschar excision and shaving were covered with autologous Meek skin graft, particulate skin graft, reticular skin graft and small skin graft respectively. The treatment outcome, CRRT time, operation times, time of recovery of serum creatinine and myoglobin, length of hospital stay, and follow-up were recorded. Results All the 4 patients were cured after transfer to our hospital. Among them, totally 5 limbs of patients No. 1 and No. 4 underwent amputation because of complication of myofascial compartment syndrome and a large amount of necrotic muscle which could not be preserved. Patients No. 1, 2, 3, and 4 were treated with CRRT for 19, 35, 14, and 25 days respectively and performed with operation for 5, 6, 10, 8 times respectively. Serum creatinine of patients No. 1, 2, 3, and 4 returned to normal on 22, 35, 37, and 48 days after transfer respectively, and their serum myoglobin returned to normal on 18, 28, 25, and 30 days after transfer respectively. Patients No. 1, 2, 3, and 4 were hospitalized for 52, 105, 148, and 156 days and discharged after basic wound healing. Follow-up for 1 to 36 months showed no abnormal renal function in 4 patients. Conclusions The early AKI in patients No. 1 and 4 was caused by rhabdomyolysis after severe burn complicated with myofascial compartment syndrome, while that of the other 2 cases were also related to hypovolemic shock and poor renal perfusion. The success rate of early AKI treatment in severely burned patients can be effectively improved by removing the causes of diseases at the same time of CRRT and actively treating burn wounds under the support of organ function and hemodynamic monitoring.
2019, 35(2): 116-124.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.007
Abstract:
Objective To investigate the role of hexokinase Ⅱ in the changes of autophagic flow in cardiomyocytes of mice with ischemia-hypoxia in vitro. Methods The hearts of totally six male and female C57BL/6 mice aged from 1 to 2 days were isolated to culture primary cardiomyocytes which were used for the following experiments. (1) The cells were divided into 6 groups according to the random number table (the same grouping method below), i. e., normal control 3, 6, and 9 h groups and ischemia-hypoxia 3, 6, and 9 h groups, with 4 wells in each group. After being regularly cultured for 48 h with Dulbecco′s modified Eagle medium/nutrient mixture F12 (DMEM/F12) medium (the same regular culture condition below), the cells in normal control 3, 6, and 9 h groups were cultured with replaced fresh DMEM/F12 medium for 3, 6, and 9 h, respectively, and the cells in ischemia-hypoxia 3, 6, and 9 h groups were cultured with replaced sugar-free serum-free medium in the low-oxygen incubator with a volume fraction of 1% oxygen and a volume fraction of 5% carbon dioxide at 37 ℃ (the same hypoxic culture condition below) for 3, 6, and 9 h, respectively. Cell viability was measured by the cell counting kit 8 (CCK-8) method. (2) The cells were grouped and treated the same as those in experiment (1), with 1 well in each group. Western blotting was used to detect the protein expressions of microtubule-associated protein 1 light chain 3 Ⅰ (LC3Ⅰ), LC3Ⅱ, p62, and hexokinase Ⅱ. (3) The cells were divided into normal control group, simple ischemia-hypoxia 9 h group, and ischemia-hypoxia 9 h+ 2-deoxyglucose (2-DG) group, with 4 wells in each group. After a regular culture for 48 h, the cells in normal control group were cultured with replaced fresh DMEM/F12 medium for 9 h; the cells in simple ischemia-hypoxia 9 h group were replaced with sugar-free serum-free medium, and the cells in ischemia-hypoxia 9 h+ 2-DG group were replaced with sugar-free serum-free medium in which 2-DG was dissolved in a concentration of 10 mmol/L (20 μmol), and then they were cultured with hypoxia for 9 h. Cell viability was measured by CCK-8 method. (4) The cells were grouped and treated the same as those in experiment (3), with 1 well in each group. Western blotting was used to detect the protein expressions of LC3Ⅰ, LC3Ⅱ, and p62. (5) The cells were grouped and treated the same as those in experiment (3), with 2 wells in each group. Transmission electron microscope was used to observe autophagosomes/autolysosomes in cardiomyocytes. (6) The cells were divided into normal control group, simple ischemia-hypoxia 9 h group, ischemia-hypoxia 9 h+ hexosinase Ⅱ small interfering RNA1 (HK-ⅡsiRNA1) group, and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group, with 4 wells in each group. The cells in normal control group and simple ischemia-hypoxia 9 h group were regularly cultured for 48 h, and the cells in ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were respectively transfected with 200 nmol/L HK-ⅡsiRNA1 and HK-ⅡsiRNA2 and then also cultured for 48 h. The cells in normal control group were cultured with replaced fresh DMEM/F12 medium for 9 h, and the cells in simple ischemia-hypoxia 9 h group, ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group, and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were cultured with replaced sugar-free serum-free medium and hypoxia for 9 h. Cell viability was measured by CCK-8 method. (7) The cells were grouped and treated the same as those in experiment (6), with 1 well in each group. Western blotting was used to detect the protein expressions of LC3Ⅰ, LC3Ⅱ, p62, and hexokinase Ⅱ. Except for experiment (5), each experiment was repeated 3 times. Data were processed with one-way analysis of variance and lest significant differencet test, and Bonferroni correction.
Results (1) The viabilities of cardiomyocytes in ischemia-hypoxia 3, 6, and 9 h groups were 0.450±0.022, 0.385±0.010, and 0.335±0.015, respectively, which were significantly lower than 0.662±0.026, 0.656±0.028, and 0.661±0.021 of the corresponding normal control 3, 6, and 9 h groups, respectively (t =6.21, 9.12, 12.48, P <0.01). (2) Compared with those of corresponding normal control 3, 6, and 9 h groups, the LC3Ⅱ/Ⅰ ratio and protein expressions of p62 and hexokinase Ⅱ in cardiomyocytes of ischemia-hypoxia 3, 6, and 9 h groups were significantly increased (t 3 h=16.15, 10.99, 5.30, t 6 h=6.79, 10.42, 9.42, t 9 h=15.76, 16.51, 7.20, P <0.05 or P <0.01). (3) The viability of cardiomyocytes in simple ischemia-hypoxia 9 h group was 0.353±0.022, which was significantly lower than 0.673±0.027 of normal control group (t =9.29, P <0.01). The viability of cardiomyocytes in ischemia-hypoxia 9 h+ 2-DG group was 0.472±0.025, which was significantly higher than that of simple ischemia-hypoxia 9 h group (t =3.60, P <0.05). (4) Compared with those of normal control group, the LC3Ⅱ/Ⅰ ratio and protein expression of p62 in cardiomyocytes of simple ischemia-hypoxia 9 h group were significantly increased (t =9.45, 8.40, P <0.01). Compared with those of simple ischemia-hypoxia 9 h group, the LC3Ⅱ/Ⅰratio and protein expression of p62 in cardiomyocytes of ischemia-hypoxia 9 h+ 2-DG group were significantly decreased (t =4.39, 4.74, P <0.05). (5) In cardiomyocytes of normal control group, only single autophagosome/autolysosome with bilayer membrane structure was observed. Compared with that of normal control group, the number of autophagosome/autolysosome with bilayer membrane structure in cardiomyocytes of simple ischemia-hypoxia 9 h group was increased significantly. Compared with that of simple ischemia-hypoxia 9 h group, the number of autophagosome/autolysosome with bilayer membrane structure in cardiomyocytes of ischemia-hypoxia 9 h+ 2-DG group was significantly decreased. (6) The viability of cardiomyocytes in simple ischemia-hypoxia 9 h group was 0.358±0.023, which was significantly lower than 0.673±0.026 in normal control group (t =9.12, P <0.01). The viabilities of cardiomyocytes in ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were 0.487±0.027 and 0.493±0.022, respectively, which were significantly higher than the viability in simple ischemia-hypoxia 9 h group (t =3.63, 4.28, P <0.05). (7) Compared with those of normal control group, the LC3Ⅱ/Ⅰratio and protein expressions of p62 and hexokinase Ⅱ in cardiomyocytes of simple ischemia-hypoxia 9 h group were significantly increased (t =6.08, 6.31, 4.83, P <0.05 or P <0.01). Compared with those of simple ischemia-hypoxia 9 h group, the LC3Ⅱ/Ⅰ ratio and protein expressions of p62 and hexokinase Ⅱ in cardiomyocytes of ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were significantly decreased (t =5.10, 7.76, 15.33, 4.17, 8.42, 12.11, P <0.05 or P <0.01).
Conclusions Ischemia-hypoxia upregulates the expression level of hexokinase Ⅱ protein in mouse cardiomyocytes cultured in vitro, which decreases the viability of cardiomyocytes by impairing autophagic flow. To inhibit the activity of hexokinase Ⅱ or its expression can alleviate the ischemia-hypoxia damage of cardiomyocytes.
Objective To investigate the role of hexokinase Ⅱ in the changes of autophagic flow in cardiomyocytes of mice with ischemia-hypoxia in vitro. Methods The hearts of totally six male and female C57BL/6 mice aged from 1 to 2 days were isolated to culture primary cardiomyocytes which were used for the following experiments. (1) The cells were divided into 6 groups according to the random number table (the same grouping method below), i. e., normal control 3, 6, and 9 h groups and ischemia-hypoxia 3, 6, and 9 h groups, with 4 wells in each group. After being regularly cultured for 48 h with Dulbecco′s modified Eagle medium/nutrient mixture F12 (DMEM/F12) medium (the same regular culture condition below), the cells in normal control 3, 6, and 9 h groups were cultured with replaced fresh DMEM/F12 medium for 3, 6, and 9 h, respectively, and the cells in ischemia-hypoxia 3, 6, and 9 h groups were cultured with replaced sugar-free serum-free medium in the low-oxygen incubator with a volume fraction of 1% oxygen and a volume fraction of 5% carbon dioxide at 37 ℃ (the same hypoxic culture condition below) for 3, 6, and 9 h, respectively. Cell viability was measured by the cell counting kit 8 (CCK-8) method. (2) The cells were grouped and treated the same as those in experiment (1), with 1 well in each group. Western blotting was used to detect the protein expressions of microtubule-associated protein 1 light chain 3 Ⅰ (LC3Ⅰ), LC3Ⅱ, p62, and hexokinase Ⅱ. (3) The cells were divided into normal control group, simple ischemia-hypoxia 9 h group, and ischemia-hypoxia 9 h+ 2-deoxyglucose (2-DG) group, with 4 wells in each group. After a regular culture for 48 h, the cells in normal control group were cultured with replaced fresh DMEM/F12 medium for 9 h; the cells in simple ischemia-hypoxia 9 h group were replaced with sugar-free serum-free medium, and the cells in ischemia-hypoxia 9 h+ 2-DG group were replaced with sugar-free serum-free medium in which 2-DG was dissolved in a concentration of 10 mmol/L (20 μmol), and then they were cultured with hypoxia for 9 h. Cell viability was measured by CCK-8 method. (4) The cells were grouped and treated the same as those in experiment (3), with 1 well in each group. Western blotting was used to detect the protein expressions of LC3Ⅰ, LC3Ⅱ, and p62. (5) The cells were grouped and treated the same as those in experiment (3), with 2 wells in each group. Transmission electron microscope was used to observe autophagosomes/autolysosomes in cardiomyocytes. (6) The cells were divided into normal control group, simple ischemia-hypoxia 9 h group, ischemia-hypoxia 9 h+ hexosinase Ⅱ small interfering RNA1 (HK-ⅡsiRNA1) group, and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group, with 4 wells in each group. The cells in normal control group and simple ischemia-hypoxia 9 h group were regularly cultured for 48 h, and the cells in ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were respectively transfected with 200 nmol/L HK-ⅡsiRNA1 and HK-ⅡsiRNA2 and then also cultured for 48 h. The cells in normal control group were cultured with replaced fresh DMEM/F12 medium for 9 h, and the cells in simple ischemia-hypoxia 9 h group, ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group, and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were cultured with replaced sugar-free serum-free medium and hypoxia for 9 h. Cell viability was measured by CCK-8 method. (7) The cells were grouped and treated the same as those in experiment (6), with 1 well in each group. Western blotting was used to detect the protein expressions of LC3Ⅰ, LC3Ⅱ, p62, and hexokinase Ⅱ. Except for experiment (5), each experiment was repeated 3 times. Data were processed with one-way analysis of variance and lest significant difference
2019, 35(2): 125-133.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.008
Abstract:
Objective To observe the effects of basic fibroblast growth factor (bFGF), hepatocyte growth factor (HGF), and vascular endothelial growth factor C (VEGF-C) on the differentiation of bone marrow mesenchymal stem cells (BMSCs) into lymphatic endothelial cells (LECs). Methods The third to the fifth passage of BMSCs of rats were collected for the following experiments. (1) BMSCs of rats were collected and divided into negative control group, CD90 group, CD44 group, and CD34 group according to the random number table (the same grouping method below), with 3 samples in each group. Phosphate buffer of 5 μL was added to cells in negative control group, and cells in the other 3 groups were added with 5 μL corresponding antibodies respectively. The positive expression of cell surface antigen was detected by flow cytometer. (2) BMSCs of rats in 3 batches were collected and divided into blank control group, VEGF-C group, HGF group, bFGF group, VEGF-C+ HGF group, VEGF-C+ bFGF group, HGF+ bFGF group, and VEGF-C+ HGF+ bFGF group, with 3 samples in each group. Cells in blank control group were added with 2 mL complete medium, cells in VEGF-C group were added with 2 mL complete medium and 10 μL VEGF-C of 10 μg/mL, cells in HGF group were added with 2 mL complete medium and 16 μL HGF of 10 μg/mL, and cells in bFGF group were added with 2 mL complete medium and 20 μL bFGF of 1 μg/mL. Cells in VEGF-C+ HGF group, VEGF-C+ bFGF group, HGF+ bFGF group, and VEGF-C+ HGF+ bFGF group were added with 2 mL complete medium and induction factors with corresponding concentration and volume as above. On 10 d of culture, the morphology of the cells was observed by the inverted phase contrast microscope, and the protein and mRNA expressions of lymphatic vessel endothelial hyaluronic acid receptor 1 (LYVE-1), VEGF receptor 3 (VEGFR3), and integrin α9 were detected by Western blotting and real-time fluorescent quantitative reverse transcription polymerase chain reaction respectively. (3) BMSCs of rats were collected and divided into blank control group, HGF+ VEGF-C+ bFGF group, bFGF+ VEGF-C+ HGF group, and VEGF-C+ HGF+ bFGF group, with 3 samples in each group. Cells in blank control group were added with 2 mL complete medium. Cells in HGF+ VEGF-C+ bFGF group were added with 2 mL complete medium, 16 μL HGF of 10 μg/mL, and 10 μL VEGF-C of 10 μg/mL, after 6 hours, 20 μL bFGF of 1 μg/mL was added. Cells in bFGF+ VEGF-C+ HGF group were added with 2 mL complete medium, 20 μL bFGF of 1 μg/mL, and 10 μL VEGF-C of 10 μg/mL, after 6 hours, 16 μL HGF of 10 μg/mL was added. Cells in VEGF-C+ HGF+ bFGF group were simultaneously added with 2 mL complete medium and the same concentration and volume of three inducing factors as above. In addition, BMSCs of rats in another 2 batches were collected and grouped, and they were dealt with the same methods as above except that the interval time of 6 hours in HGF+ VEGF-C+ bFGF group and bFGF+ VEGF-C+ HGF group was adjusted to 12 and 24 hours. On 10 d of culture, protein expressions of LYVE-1, VEGFR3, and integrin α9 were detected by Western blotting. Data were processed with analysis of variance of factorial design, one-way analysis of variance, and least significant differencet test, and Bonferroni correction.
Results (1) The positive expression rates of surface antigen of cells in negative control group, CD90 group, CD44 group, and CD34 group were 0.39%, 99.84%, 99.90%, and 0.57%, respectively. (2) On 10 d of culture, cells in blank control group, HGF group, bFGF group, and HGF+ bFGF group presented long fusiform, while cells in the other groups presented polygonal shape. (3) On 10 d of culture, there were no protein expressions of LYVE-1, VEGFR3, and integrin α9 in cells of blank control group, HGF group, bFGF group, and HGF+ bFGF group. On 10 d of culture, protein expressions of LYVE-1, VEGFR3, and integrin α9 in cells of VEGF-C+ HGF+ bFGF group were significantly higher than those in VEGF-C group (t =24.21, 11.04, 15.43, P <0.01), VEGF-C+ HGF group (t =10.81, 9.93, 10.20, P <0.01), and VEGF-C+ bFGF group (t =11.67, 6.32, 19.00, P <0.01). Protein expressions of LYVE-1 in cells of VEGF-C+ HGF group and VEGF-C+ bFGF group were significantly higher than the protein expression in VEGF-C group (t =8.69, 15.20, P <0.01). Protein expression of VEGFR3 in cells of VEGF-C+ bFGF group was obviously higher than the protein expressions in VEGF-C group and VEGF-C+ HGF group (t =8.67, 7.21, P <0.01). Protein expression of integrin α9 in cells of VEGF-C+ HGF group was obviously higher than the protein expressions in VEGF-C group and VEGF-C+ bFGF group (t =8.80, 8.83, P <0.01). (4) On 10 d of culture, there were no mRNA expressions of LYVE-1, VEGFR3, and integrin α9 in cells of blank control group, HGF group, bFGF group, and HGF+ bFGF group. On 10 d of culture, mRNA expressions of LYVE-1 and VEGFR3 in cells of VEGF-C group were significantly lower than those in VEGF-C+ bFGF group and VEGF-C+ HGF+ bFGF group (t LYVE-1=6.22, 18.01, t VEGFR3=8.49, 15.34, P <0.01), and mRNA expression of integrin α9 were significantly lower than that in VEGF-C+ HGF group and VEGF-C+ HGF+ bFGF group (t =13.24, 9.65, P <0.01). The mRNA expressions of LYVE-1, VEGFR3, and integrin α9 in cells of VEGF-C+ HGF+ bFGF group were obviously higher than those in VEGF-C+ HGF group and VEGF-C+ bFGF group (t =13.92, 11.95, 13.72, 5.27, 5.64, 9.10, P <0.01). Compared with those of VEGF-C+ bFGF group, the mRNA expression of VEGFR3 of cells in VEGF-C+ HGF group was significantly lower (t =6.91, P <0.01), while the mRNA expression of integrin α9 of cells in VEGF-C+ HGF group was significantly higher (t =11.69, P <0.01). (5) On 10 d of culture at interval time of 6, 12, 24 h, there were no protein expressions of LYVE-1, VEGFR3, or integrin α9 in cells of blank control group. On 10 d of culture at interval time of 6, 12, 24 h, the protein expressions of LYVE-1, VEGFR3, and integrin α9 in cells of HGF+ VEGF-C+ bFGF group, bFGF+ VEGF-C+ HGF group, and VEGF-C+ HGF+ bFGF group were close (F 6 h=2.25, 2.47, 2.19, F 12 h=2.93, 1.47, 3.25, F 24 h=0.28, 0.20, 1.01, P >0.05).
Conclusions VEGF-C is a necessary factor for inducing BMSCs to differentiate into LECs. HGF and bFGF may promote the differentiation by up-regulating the expressions of integrin α9 and VEGFR3 respectively. But the induction effects of the two factors may be independent. The combination of VEGF-C, HGF, and bFGF have the best effects of promoting differentiation.
Objective To observe the effects of basic fibroblast growth factor (bFGF), hepatocyte growth factor (HGF), and vascular endothelial growth factor C (VEGF-C) on the differentiation of bone marrow mesenchymal stem cells (BMSCs) into lymphatic endothelial cells (LECs). Methods The third to the fifth passage of BMSCs of rats were collected for the following experiments. (1) BMSCs of rats were collected and divided into negative control group, CD90 group, CD44 group, and CD34 group according to the random number table (the same grouping method below), with 3 samples in each group. Phosphate buffer of 5 μL was added to cells in negative control group, and cells in the other 3 groups were added with 5 μL corresponding antibodies respectively. The positive expression of cell surface antigen was detected by flow cytometer. (2) BMSCs of rats in 3 batches were collected and divided into blank control group, VEGF-C group, HGF group, bFGF group, VEGF-C+ HGF group, VEGF-C+ bFGF group, HGF+ bFGF group, and VEGF-C+ HGF+ bFGF group, with 3 samples in each group. Cells in blank control group were added with 2 mL complete medium, cells in VEGF-C group were added with 2 mL complete medium and 10 μL VEGF-C of 10 μg/mL, cells in HGF group were added with 2 mL complete medium and 16 μL HGF of 10 μg/mL, and cells in bFGF group were added with 2 mL complete medium and 20 μL bFGF of 1 μg/mL. Cells in VEGF-C+ HGF group, VEGF-C+ bFGF group, HGF+ bFGF group, and VEGF-C+ HGF+ bFGF group were added with 2 mL complete medium and induction factors with corresponding concentration and volume as above. On 10 d of culture, the morphology of the cells was observed by the inverted phase contrast microscope, and the protein and mRNA expressions of lymphatic vessel endothelial hyaluronic acid receptor 1 (LYVE-1), VEGF receptor 3 (VEGFR3), and integrin α9 were detected by Western blotting and real-time fluorescent quantitative reverse transcription polymerase chain reaction respectively. (3) BMSCs of rats were collected and divided into blank control group, HGF+ VEGF-C+ bFGF group, bFGF+ VEGF-C+ HGF group, and VEGF-C+ HGF+ bFGF group, with 3 samples in each group. Cells in blank control group were added with 2 mL complete medium. Cells in HGF+ VEGF-C+ bFGF group were added with 2 mL complete medium, 16 μL HGF of 10 μg/mL, and 10 μL VEGF-C of 10 μg/mL, after 6 hours, 20 μL bFGF of 1 μg/mL was added. Cells in bFGF+ VEGF-C+ HGF group were added with 2 mL complete medium, 20 μL bFGF of 1 μg/mL, and 10 μL VEGF-C of 10 μg/mL, after 6 hours, 16 μL HGF of 10 μg/mL was added. Cells in VEGF-C+ HGF+ bFGF group were simultaneously added with 2 mL complete medium and the same concentration and volume of three inducing factors as above. In addition, BMSCs of rats in another 2 batches were collected and grouped, and they were dealt with the same methods as above except that the interval time of 6 hours in HGF+ VEGF-C+ bFGF group and bFGF+ VEGF-C+ HGF group was adjusted to 12 and 24 hours. On 10 d of culture, protein expressions of LYVE-1, VEGFR3, and integrin α9 were detected by Western blotting. Data were processed with analysis of variance of factorial design, one-way analysis of variance, and least significant difference
Chen Wei,
Wei Zairong,
Wu Bihua,
Yang Chenglan,
Jin Wenhu,
Gong Feiyu,
Sun Guangfeng,
Nie Kaiyu,
Wang Dali
2019, 35(2): 134-142.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.009
Abstract:
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 1×105 cells/mL were co-cultured as Schwann cells+ Fbs co-culture group, and 2 mL Schwann cells in the concentration of 1×105 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 (2×106 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 2×106 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.
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 (
2019, 35(2): 143-147.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.010
Abstract:
Objective To investigate and analyze the actual intake of protein and energy in adult patients with severe burns during post burn days (PBDs) 3 to 14. Methods Records of 52 adult patients with severe burns [37 males and 15 females, (37±9) years old], admitted to the Department of Plastic Surgery and Burns of Tianjin First Central Hospital from January 1st 2011 to December 31st 2017 and meeting the study inclusion criteria, were retrospectively analyzed. Nutrition intake from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations of patients during PBDs 3 to 14 were obtained from critical care records. During PBDs 3 to 7 and PBDs 8 to 14, the personal daily total energy intake and the ratio of it to energy target of patients were calculated and compared; the personal daily intake of carbohydrate, fat, and protein and calorigenic percentages of carbohydrate, fat, and protein accounted for total energy intake, and the ratios of non-protein calories to total nitrogen of patients were calculated and compared; the personal daily energy and protein intake of patients from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations were analyzed; the percentages of energy intake from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations accounted for total energy intake, and the percentages of protein intake from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations accounted for total protein intake of patients were calculated. Vomiting and diarrhea of patients during PBDs 3 to 7 and PBDs 8 to 14 were recorded. Levels of serum albumin, prealbumin, blood glucose, and triglycerides, 24-hour excretion of urinary nitrogen, nitrogen balance values of patients on PBDs 7 and 14 were recorded or calculated. Data were processed with pairedt test and chi-square test.
Results (1) The personal daily total energy intake of patients during PBDs 3 to 7 and PBDs 8 to 14 were (8 696±573) and (11 980±1 259) kJ respectively, and ratios of them to energy target [(13 290±1 561) kJ] were 65.4% and 90.1% respectively. The personal daily total energy intake of patients during PBDs 3 to 7 was obviously lower than that during PBDs 8 to 14 (t =18.172, P <0.01). (2) The personal daily intake of carbohydrate, fat, and protein of patients during PBDs 8 to 14 were obviously higher than those during PBDs 3 to 7 (t =15.628, 22.231, 10.403, P <0.01). The personal daily calorigenic percentages of carbohydrate, fat, and protein accounted for total energy intake of patients were 56.8%, 25.1%, and 18.3% respectively during PBDs 3 to 7 and 54.2%, 27.0%, and 18.7% respectively during PBDs 8 to 14. The calorigenic constituent ratio of personal daily intake of carbohydrate, fat, and protein accounted for total energy intake of patients during PBDs 3 to 7 was close to that during PBDs 8 to 14 (χ 2=0.185, P >0.05). The ratios of non-protein calories to total nitrogen (kJ∶g) of patients during PBDs 3 to 7 and PBDs 8 to 14 were 469∶ 1 and 456∶ 1 respectively. (3) The personal daily energy intake of patients from routes of oral diet and parenteral nutrition preparations during PBDs 8 to 14 [(4 394±978), (5 723±898) kJ] were obviously higher than those during PBDs 3 to 7 [(2 137±453), (4 855±825) kJ, t =26.516, 6.583, P <0.01], while the personal daily energy intake of patients from routes of enteral nutrition preparations during PBDs 8 to 14 was close to that during PBDs 3 to 7 (t =1.922, P >0.05). The constituent ratio of personal daily energy during PBDs 3 to 7 was close to that during PBDs 8 to 14 (χ 2=4.100, P >0.05). The personal daily protein intake of patients from route of oral diet during PBDs 8 to 14 was (58±22) g, obviously higher than (25±6) g during PBDs 3 to 7 (t =14.514, P <0.01). The personal daily protein intake of patients from routes of enteral nutrition preparations and parenteral nutrition preparations during PBDs 8 to 14 was close to those during PBDs 3 to 7 (t =1.924, 1.110, P >0.05). The constituent ratio of personal daily protein intake from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations accounted for total protein intake during PBDs 8 to 14 was close to that during PBDs 3 to 7 (χ 2=5.634, P >0.05). (4) There were 3 patients with vomiting and 4 patients with diarrhea during PBDs 3 to 7, and 1 patient experienced both of them during PBDs 8 to 14. The levels of serum albumin, prealbumin, blood glucose, and triglycerides, 24-hour excretion of urinary nitrogen, and nitrogen balance values of patients on PBDs 7 and 14 were (29±4) and (30±4) g/L, (132±42) and (171±48) mg/L, (7.4±2.8) and (6.7±2.8) mmol/L, (1.5±0.7) and (1.4±0.7) mmol/L, (30.5±4.3) and (34.5±2.2) g, -(25.1±2.6) and -(23.7±3.9) g, respectively.
Conclusions The personal daily total energy intake of patients during PBDs 3 to 7 was lower than that during PBDs 8 to 14. The calorigenic constituent ratio of personal daily intake of carbohydrate, fat, and protein accounted for total energy of patients during PBDs 3 to 7 was close to that during PBDs 8 to 14. Energy and protein intake were mostly derived from parenteral nutrition preparations during PBDs 3 to 7, while those during PBDs 8 to 14 were mainly derived from parenteral nutrition preparations and oral diet.
Objective To investigate and analyze the actual intake of protein and energy in adult patients with severe burns during post burn days (PBDs) 3 to 14. Methods Records of 52 adult patients with severe burns [37 males and 15 females, (37±9) years old], admitted to the Department of Plastic Surgery and Burns of Tianjin First Central Hospital from January 1st 2011 to December 31st 2017 and meeting the study inclusion criteria, were retrospectively analyzed. Nutrition intake from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations of patients during PBDs 3 to 14 were obtained from critical care records. During PBDs 3 to 7 and PBDs 8 to 14, the personal daily total energy intake and the ratio of it to energy target of patients were calculated and compared; the personal daily intake of carbohydrate, fat, and protein and calorigenic percentages of carbohydrate, fat, and protein accounted for total energy intake, and the ratios of non-protein calories to total nitrogen of patients were calculated and compared; the personal daily energy and protein intake of patients from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations were analyzed; the percentages of energy intake from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations accounted for total energy intake, and the percentages of protein intake from routes of oral diet, enteral nutrition preparations, and parenteral nutrition preparations accounted for total protein intake of patients were calculated. Vomiting and diarrhea of patients during PBDs 3 to 7 and PBDs 8 to 14 were recorded. Levels of serum albumin, prealbumin, blood glucose, and triglycerides, 24-hour excretion of urinary nitrogen, nitrogen balance values of patients on PBDs 7 and 14 were recorded or calculated. Data were processed with paired
Zhou Qin,
Li Shuangshuang,
Wang Qing,
Lu Ying,
Si Yanning,
Wang Lina,
Zhao Deli,
Luo Xufang,
Hu Xuehui
2019, 35(2): 148-152.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.011
Abstract:
Objective To explore the influence of cluster nursing intervention on inadequate drainage in vacuum sealing drainage (VSD) for inpatients in burn unit. Methods From October to December 2016, 60 patients, aged (43.6±2.8) years admitted to our department, receiving VSD treatment and conforming to the inclusion criteria, were included in the routine nursing group, and among the patients, 37 cases were male and 23 cases were female. From May to July 2017, 58 patients, aged (44.2±3.2) years admitted to our department, receiving VSD treatment and conforming to the inclusion criteria, were included in the cluster nursing group, and among the patients, 36 cases were male and 22 cases were female. The patients′ medical records were retrospectively analyzed. After VSD treatment, patients in routine nursing group received routine nursing, and patients in cluster nursing group received cluster nursing. A cluster intervention group was formed and headed by a chief surgeon. The cluster nursing plan was formulated and implemented strictly from the following six aspects of material preparation, negative pressure value control and negative pressure mode setting, drainage tube nursing, semi-permeable membrane reinforcement, standardizing changing process and timing of drainage capsule, and health education. During VSD treatment, the incidence of inadequate drainage, reasons of inadequate drainage and the occurrences, occurrences of inadequate drainage of wounds in different types and sites, and satisfaction of patients in two groups were observed and calculated. The patient satisfaction items included procedure of drainage capsule replacement, the method of tube fixation, the content and form of health education. Data were processed with independent samplet test and chi-square test.
Results (1) During VSD treatment, the incidence of inadequate drainage of patients in routine nursing group was 43.33% (130/300), which was significantly higher than 17.24% (50/290) in cluster nursing group (χ 2=43.350, P <0.01). (2) During VSD treatment, the incidences of inadequate drainage caused by blockage of drainage tube due to scabbing of drainage, low negative pressure, air leakage of semi-permeable membrane, improper changing process of drainage capsule, shedding, compression, reversal of drainage tube of patients in cluster nursing group were 7.93% (23/290), 4.48% (13/290), 1.72% (5/290), 1.03% (3/290), and 2.07% (6/290), respectively, significantly lower than 16.67% (50/300), 11.67% (35/300), 4.33% (13/300), 4.00% (12/300), and 6.67% (20/300) in routine nursing group (χ 2=10.379, 22.951, 4.832, 7.840, 7.399, P <0.05 or P <0.01). (3) During VSD treatment, the incidences of inadequate drainage of burn wounds, trauma wounds, pressure ulcer, venous ulcer in lower limbs, and diabetic foot of patients in cluster nursing group were significantly lower than those in routine nursing group (χ 2=17.835, 6.809, 9.478, 4.939, 8.631, P <0.05 or P <0.01). During VSD treatment, the incidences of inadequate drainage of wounds in different types of patients in the same group were close (χ 2=0.434, 0.057, P >0.05). (4) During VSD treatment, the incidences of inadequate drainage of wounds in limbs, trunk, buttocks, and sacrococcyx of patients in cluster nursing group were significantly lower than those in routine nursing group (χ 2=31.892, 9.588, 4.939, 4.549, P <0.05 or P <0.01). During VSD treatment, the incidences of inadequate drainage of wounds in different wound sites of patients in the same group were close (χ 2=0.071, 0.069, P >0.05). (5) The satisfaction scores in changing process of drainage capsule, method of tube fixation, content and form of health education of patients in cluster nursing group after VSD treatment were significantly higher than those in routine nursing group (t =5.166, 4.471, 7.958, 8.975, P <0.01).
Conclusions Cluster nursing intervention on patients receiving VSD treatment could reduce the incidences of inadequate drainage of wounds in different types and sites caused by various reasons. It also can improve patient satisfaction.
Objective To explore the influence of cluster nursing intervention on inadequate drainage in vacuum sealing drainage (VSD) for inpatients in burn unit. Methods From October to December 2016, 60 patients, aged (43.6±2.8) years admitted to our department, receiving VSD treatment and conforming to the inclusion criteria, were included in the routine nursing group, and among the patients, 37 cases were male and 23 cases were female. From May to July 2017, 58 patients, aged (44.2±3.2) years admitted to our department, receiving VSD treatment and conforming to the inclusion criteria, were included in the cluster nursing group, and among the patients, 36 cases were male and 22 cases were female. The patients′ medical records were retrospectively analyzed. After VSD treatment, patients in routine nursing group received routine nursing, and patients in cluster nursing group received cluster nursing. A cluster intervention group was formed and headed by a chief surgeon. The cluster nursing plan was formulated and implemented strictly from the following six aspects of material preparation, negative pressure value control and negative pressure mode setting, drainage tube nursing, semi-permeable membrane reinforcement, standardizing changing process and timing of drainage capsule, and health education. During VSD treatment, the incidence of inadequate drainage, reasons of inadequate drainage and the occurrences, occurrences of inadequate drainage of wounds in different types and sites, and satisfaction of patients in two groups were observed and calculated. The patient satisfaction items included procedure of drainage capsule replacement, the method of tube fixation, the content and form of health education. Data were processed with independent sample
2019, 35(2): 153-156.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.012
Abstract:
Pressure ulcer is localized damage to the skin and subcutaneous tissue usually over a bony prominence as a result of prolonged pressure, shear force, and friction. The effect of external force on soft tissue is affected by many factors, such as local microenvironment, tissue blood supply, nutritional status, and underlying diseases of patients. Although great efforts have been made by mankind to prevent and treat pressure ulcer in last decades, its prevalence is still high, and the curative effect is still not impressive. The treatment of pressure ulcer is a challenge today, and it is necessary to seek new treatment methods. However, the prerequisite for exploring new treatment methods is to find a proper animal model and further explore new therapies through animal experiments. The pathogenesis of pressure ulcer is complex, and the formation process is affected by a variety of factors. To date, there is no recognized standard animal model. We review the pathogenesis of pressure ulcer and the recently reported animal models of pressure ulcer, so as to provide basic experimental basis for further research on the occurrence, development, and prevention of pressure ulcer.
Pressure ulcer is localized damage to the skin and subcutaneous tissue usually over a bony prominence as a result of prolonged pressure, shear force, and friction. The effect of external force on soft tissue is affected by many factors, such as local microenvironment, tissue blood supply, nutritional status, and underlying diseases of patients. Although great efforts have been made by mankind to prevent and treat pressure ulcer in last decades, its prevalence is still high, and the curative effect is still not impressive. The treatment of pressure ulcer is a challenge today, and it is necessary to seek new treatment methods. However, the prerequisite for exploring new treatment methods is to find a proper animal model and further explore new therapies through animal experiments. The pathogenesis of pressure ulcer is complex, and the formation process is affected by a variety of factors. To date, there is no recognized standard animal model. We review the pathogenesis of pressure ulcer and the recently reported animal models of pressure ulcer, so as to provide basic experimental basis for further research on the occurrence, development, and prevention of pressure ulcer.
2019, 35(2): 157-160.
doi: 10.3760/cma.j.issn.1009-2587.2019.02.013
Abstract:
As a natural extract from turmeric, curcumin has extensive pharmacological effects, such as anti-tumor, anti-inflammation, anti-oxidative stress, anti-microbial, immunoregulation and so on. In recent years, an increasing number of basic and clinical researches have shown that curcumin takes therapeutic effects on various diseases, such as gastrointestinal diseases, cardiovascular diseases, autoimmune diseases, neuropsychiatric diseases and so on. Many of the pharmacological effects and mechanisms of curcumin are associated with protective effects of intestinal mucosal barrier. It can protect intestinal mucosal barrier through mutiple pathways, including anti-inflammation, anti-oxidative stress, anti-bacterial, anti-apoptosis, regulating intestinal microecology and intestinal immune response and so on. This paper summarizes the protective effects of curcumin on intestinal barrier function and the mechanism, in order to provide new ideas for diagnosis and treatment of intestinal dysfunction.
As a natural extract from turmeric, curcumin has extensive pharmacological effects, such as anti-tumor, anti-inflammation, anti-oxidative stress, anti-microbial, immunoregulation and so on. In recent years, an increasing number of basic and clinical researches have shown that curcumin takes therapeutic effects on various diseases, such as gastrointestinal diseases, cardiovascular diseases, autoimmune diseases, neuropsychiatric diseases and so on. Many of the pharmacological effects and mechanisms of curcumin are associated with protective effects of intestinal mucosal barrier. It can protect intestinal mucosal barrier through mutiple pathways, including anti-inflammation, anti-oxidative stress, anti-bacterial, anti-apoptosis, regulating intestinal microecology and intestinal immune response and so on. This paper summarizes the protective effects of curcumin on intestinal barrier function and the mechanism, in order to provide new ideas for diagnosis and treatment of intestinal dysfunction.
