Effects of insulin therapy on skeletal muscle wasting in severely scalded rats and its related mechanism / 中华烧伤杂志

Objective@#To explore the effects of insulin therapy on skeletal muscle wasting (SMW) in severely scalded rats and its related mechanism.@*Methods@#Totally 48 male Wistar rats aged 7-8 weeks were divided into simple scald (SS) group and insulin therapy (IT) group according to the random number table, with 24 rats in each group. After weighing the body mass and measuring the blood glycemic level of the tail end with a glucometer, the rats in the two groups were immersed in hot water at 94 ℃ for 12 seconds to make a full-thickness dorsal scald model involving 30% total body surface area. Rats in group IT were subcutaneously injected with 1 U/kg insulin glargine at 8: 00 a day from post injury day (PID) 1 to 7, whilst rats in group SS were given the same amount of normal saline. Rats in the two groups were given 10 mL/kg enteral nutritional emulsion by intragastric infusion at 8: 00 (after insulin administration), 13: 00, and 18: 00 a day respectively from PID 1 to 7. The blood glycemic levels of tail end of rats in the two groups were measured by glucometer before insulin administration on PID 1-4, 6, and 7 and on every morning of PID 8, 9, 11, 12, and 14. The body mass of rats in the two groups on PID 14 without any treatment was weighed. Eight rats from each group were collected respectively on PID 4, 7, and 14 to harvest tibialis anterior muscle (TAM) samples. The mass of TAM on PID 14 was weighed. The ultrastructural changes of TAM myocytes on PID 7 were observed with transmission electron microscope. The apoptotic rates of TAM myocytes on PID 4, 7, and 14 were assessed by the assay of terminal deoxynucleotidyl transferase-mediated deoxyuridinetriphate-biotin nick end labeling, the expressions of cysteine-aspartic protease-3 (caspase-3) of TAM on PID 4, 7, and 14 were detected with immunohistochemistry, and protein expressions of endoplasmic reticulum (ER) stress (ERS) associated proteins glucose-regulated protein 78 (GRP78), CCAAT/enhancer binding protein-homologous protein (CHOP), and activated caspase-12 of TAM on PID 4, 7, and 14 were detected with Western blotting. Data were processed with completely random design t test, analysis of variance for repeated measurement, analysis of variance for factorial design, t test, and Bonferroni correction.@*Results@#The blood glycemic level and body mass of rats in the two groups before injury were similar (t=0.204, 0.405, P>0.05). There were no statistically significant differences in blood glycemic levels of rats between the two groups on PID 1, 6, 9, 11, 12, and 14 (t=0.229, 3.339, 1.610, 0.178, 0.181, 0.079, P>0.05). Compared with those of group SS, blood glycemic levels of rats in group IT were significantly lower on PID 2, 3, 4, 7, and 8 (t=7.245, 4.165, 4.609, 4.018, 3.995, P<0.05 or P<0.01). On PID 14, the body mass and TAM mass of rats in group IT were (271±19) g and (0.47±0.05) g respectively, both obviously higher than (254±12) g and (0.43±0.04) g of group SS (t=2.159, 2.375, P<0.05). On PID 7, nuclear pyknosis and deformation, chromosome misdistribution, and ER swelling in TAM myocytes of rats in group SS were observed; the apoptotic alterations and ER swelling of TAM myocytes were alleviated in rats of group IT as compared with those of group SS. The apoptotic rates of TAM myocytes of rats in group IT were obviously lower than those of group SS on PID 4, 7, and 14 (t=4.262, 9.153, 9.799, P<0.01). The expressions of caspase-3 in TAM of rats in group IT were obviously lower than those of group SS on PID 7 and 14 (t=10.429, 7.617, P<0.01). Compared with those of group SS, the protein expressions of GRP78 were obviously increased on PID 4 and 14 (t=4.172, 4.437, P<0.05), the protein expressions of activated caspase-12 were obviously decreased on PID 7 and 14 (t=11.049, 11.181, P<0.01), and the protein expressions of CHOP were obviously decreased on PID 4, 7, and 14 (t=13.837, 9.572, 6.930, P<0.01) in TAM of rats in group IT.@*Conclusions@#Insulin therapy may reduce skeletal muscle myocytes apoptosis and SMW by alleviating ERS in rats with severe scald.

Effects of platelet-rich plasma combined with polylactic acid/polycaprolactone on healing of pig deep soft tissue defect caused by fragment injury / 中华烧伤杂志

Objective@#To investigate the effects of platelet-rich plasma (PRP) combined with polylactic acid/polycaprolactone (PLA/PCL) on healing of mininature pig deep soft tissue defect caused by fragment injury.@*Methods@#Two male Bama miniature pigs with 11 to 12 months (the same below) were selected by lottery to prepare PRP. The other twenty-seven male Bama miniature pigs were used to reproduce deep soft tissue defect caused by high-explosive ammunition fragment injury on bilateral posterior femoral region. According to the random number table, 27 pigs were divided into control group, material group, and PRP+material group, with 9 pigs in each group. After debridement, wounds of pigs in material group and PRP+material group were filled with PLA/PCL and PLA/PCL+2 mL activated PRP, respectively. Pigs in each group received suture of full-thickness skin to close the wounds. The operative duration was recorded. The length and volume of wounds of pigs in the above groups were measured immediately after surgery. In 1, 2, and 4 weeks after surgery, 3 pigs in each group were sacrificed to collect femoral wounds tissue on two sides, and PLA/PCL were collected from wounds of pigs in material group and PRP+material group for general observation of wounds tissue and degradation of the material. In 2 and 4 weeks after surgery, wounds tissue was obtained to observe the histological changes by hematoxylin-eosin staining, and expressions of transforming growth factor β (TGF-β) and vascular endothelial growth factor (VEGF), and angiogenesis were determined by immunohistochemical method. In 1, 2, and 4 weeks after surgery, wounds tissue was collected to determine mRNA expressions of TGF-β and VEGF by real-time quantitative reverse transcription polymerase chain reaction. Data were processed with analysis of variance of factorial design, one-way analysis of variance, and least significant difference-t test.@*Results@#(1) There were no significantly statistical differences in length and volume of the wounds of pigs among the three groups (F=0.336, 0.282, P>0.05). The operative duration in control group [(30.9±2.1)min] was significantly shorter than that of material group [(39.7±2.2)min] and PRP+material group[(40.0±2.6)min], t=-11.45, -11.88, P<0.01. (2) There were respectively 10, 7, and 5 wounds tissue with infection in pigs of control group, material group, and PRP+material group. In 1, 2, 4 weeks after surgery, all of the wounds tissue of pigs was infected in control group, while none of wounds tissue of pigs was infected in material group and PRP+material group. In pigs of material group and PRP+material group, materials and tissue were easily separated in 1 week after surgery; some materials were integrated with tissue and showed a tendency of degradation in 2 weeks after surgery; materials were completely embedded with tissue in 4 weeks after surgery. (3) In pigs of control group, erythrocytes and inflammatory cells infiltration in wounds tissue were observed in 2 weeks after surgery, and necrotic tissue and inflammatory cells infiltration in wounds tissue were still observed in 4 weeks after surgery. In pigs of material group and PRP+material group, a large number of erythrocytes and inflammatory cells infiltration were observed in 2 weeks after surgery. Compared with that of material group, wounds tissue of pigs in PRP+material group had no inflammatory cells infiltration in 4 weeks after surgery. (4) Protein expressions of TGF-β in fibroblasts and multinuclear macrophagocytes, VEGF in fibroblasts and vascular endothelial cells, and blood vessel formation in wounds tissue of pigs in PRP+material group were significantly more than those of pigs in control group and material group in 2 and 4 weeks after surgery. (5) The mRNA expression of TGF-β in wounds tissue of pigs in material group was significantly higher than that in control group in 4 weeks after surgery (t=-3.93, P<0.01). Compared with those of pigs in control group and material group, the mRNA expression of TGF-β in wounds tissue of pigs in PRP+material group was significantly increased at each time point (t=9.23, 13.81, 11.73, -7.51, -12.04, -7.80, P<0.01). The mRNA expression of VEGF in wounds tissue of pigs increased significantly in material group compared with that of pigs in control group in 4 weeks after surgery (t=-3.94, P<0.01). Compared with those of pigs in control group and material group, the mRNA expression of VEGF in wounds tissue increased significantly in wound tissue of pigs in PRP+material group at each time point (t=12.33, 3.95, 7.97, -11.36, -2.97, -4.04, P<0.01).@*Conclusions@#PRP combined with PLA/PCL can accelerate wound healing of deep soft tissue defect of mininature pigs caused by fragment injury by providing physical scaffold for newborn tissue growth, promoting mRNA and protein expressions of TGF-β and VEGF.

Curative effects of platelet-rich plasma combined with negative-pressure wound therapy on sternal osteomyelitis and sinus tract after thoracotomy / 中华烧伤杂志

<p><b>OBJECTIVE</b>To observe the curative effects of platelet-rich plasma (PRP) combined with negative-pressure wound therapy (NPWT) on patients with sternal osteomyelitis and sinus tract after thoracotomy.</p><p><b>METHODS</b>Sixty-two patients with sternal osteomyelitis and sinus tract after thoracotomy, hospitalized from March 2011 to June 2015, were retrospectively analyzed. Based on whether receiving PRP or not, patients were divided into two groups, group NPWT ( 22 patients hospitalized from March 2011 to December 2012) and combination treatment group (CT, 40 patients hospitalized from January 2013 to June 2015). After debridement, patients in group NPWT were treated with continuous NPWT (negative pressure values from -15.96 to -13.30 kPa), while those in group CT were treated with PRP gel (blood platelet counts in PRP ranged from 1 450×10(9)/L to 1 800×10(9)/L, with 10-15 mL in each dosage) made on the surgery day to fill the sinus tract and wound, followed by NPWT. Negative pressure materials were changed every 5 days until 20 days after surgery in patients of both groups. PRP gel was replenished before changing of negative pressure materials in patients of group CT. The sinus tract sealing time, wound healing time, number of patients who had secondary repair surgery, number of patients who had recurrence of sinus tract within three months after wound healing, and length of hospital stay were recorded. Data were processed with t test, Fisher's exact test, and chi-square test.</p><p><b>RESULTS</b>The sinus tract sealing time, wound healing time, and length of hospital stay in patients of group CT were (16±8), (27±13), and (43±13) d respectively, which were all significantly shorter than those in group NPWT [(29±14), (41±17), and (60±20) d, with t values from 3.88 to 4.67, P values below 0.01]. The number of patients who had secondary repair surgery in group CT was less than that in group NPWT (P<0.01). There was no statistically significant difference in the number of patients who had recurrence of sinus tract between two groups (P>0.05).</p><p><b>CONCLUSIONS</b>Compared with NPWT only, PRP combined with NPWT has great curative effects on patients with sternal osteomyelitis and sinus tract after thoracotomy, for it shortens sinus tract sealing time, wound healing time, and length of hospital stay, and avoids the secondary repair surgery. This method is simple and safe with little injury.</p>

Effects of endotoxin/lipopolysaccharide on proliferation and apoptosis of human umbilical cord mesenchymal stem cells / 中华烧伤杂志

<p><b>OBJECTIVE</b>To investigate the effects of different concentrations of lipopolysaccharide (LPS) on proliferation and apoptosis of human umbilical cord mesenchymal stem cells (hUCMSCs) in vitro, and to explore their possible mechanism.</p><p><b>METHODS</b>hUCMSCs from umbilical cord tissue of full-term healthy fetus delivered by caesarean section were isolated and cultured in vitro using tissue attachment method. The 3rd passage hUCMSCs were used in the study. Cells were divided into groups A, B, C, D, and E, which were treated with DMEM/F12 medium containing 0, 0.1, 1.0, 10.0, and 100.0 µg/mL of LPS respectively. In groups B, C, D, and E, methyl-thiazole-tetrazolium assay was used to detect proliferative activity of hUCMSCs at post treatment hour (PTH) 12, 24, and 48 (denoted as absorption value), with 5 samples in each group at each time point; apoptosis of hUCMSCs at PBH 24 was identified with acridine orange-ethidium bromide (AO-EB) staining, with 4 samples in each group; apoptotic rate of hUCMSCs was determined by flow cytometer, with 5 samples in each group. Above-mentioned indexes were determined in group A at the same time points. Data were processed with analysis of variance and LSD- t test.</p><p><b>RESULTS</b>(1) There was no statistically significant difference in proliferative activity of hUCMSCs at PTH 12 among groups A, B, C, D, and E (with t values from -1.67 to 1.33, P values above 0.05). Compared with that of group A, proliferative activity of hUCMSCs was increased in groups B, C, and D at PTH 24 and 48 (with t values from -13.42 to 17.34, P < 0.05 or P < 0.01), especially so in group C. Proliferative activity of hUCMSCs was lower in group E at PTH 24 and 48 than in group A (with t values respectively 8.64 and 17.34, P values below 0.01). (2) Obvious apoptosis of hUCMSCs was observed in group E but not in the other 4 groups with AO-EB staining. (3) Apoptosis rates of hUCMSCs in groups A, B, C, D, and E were respectively (3.1 ± 0.6)%, (2.6 ± 0.7)%, (2.9 ± 0.8)%, (3.1 ± 0.4)%, (25.1 ± 2.7)% (F = 272.19, P < 0.01). Apoptotic rate of hUCMSCs in group B, C, or D was respectively close to that in group A (with t values respectively 1.22, 0.57, -0.14, P values above 0.05), but it was higher in group E than in group A (t = -17.63, P < 0.01).</p><p><b>CONCLUSIONS</b>hUCMSCs proliferation may be promoted by low concentration of LPS. hUCMSCs proliferation is inhibited or induced to apoptosis along with the increase in concentration of LPS, and it may be related to activation of different major molecular signaling pathways by different concentrations of LPS.</p>

Protective effects of growth hormone on gastric mucosa of rats with hemorrhagic shock / 解放军医学杂志

Objective To investigate the protective effect of recombinant human growth hormone (rhGH) on gastric mucosa of rats with hemorrhagic shock. Methods Eighteen healthy male Sprague Dawley (SD) rats were randomly assigned into 3 groups, 6 rats for each group: control group, hemorrhagic shock group and rhGH treated group. The animals in control group were subjected to anaesthesia and intubation, without hemorrhagic shock and resuscitation, and all the parameters were determined 4h after intubation. The animals in hemorrhagic shock group and the rhGH treated group were not only given anaesthesia, intubation for carotid artery and internal jugular vein, but also reproduced as hemorrhagic shock-resuscitation model by bleeding from carotid artery and blood transfusion via internal jugular vein. The animals in rhGH treated group were given rhGH (1.5U/kg) when resuscitation began. In the hemorrhagic shock group and rhGH treated group all the parameters were determined 2h after resuscitation. Gastric mucosal blood flow (GMBF) was determined with laser doppler flowmetry (LDF), and the extent of gastric mucosal injury was assessed with optical microscope and transmission electron microscope (TEM). Results GMBF of the rats in hemorrhagic shock group was significantly lower than that in control group (260.4?49.6bpu vs. 418.6?57.3bpu, P0.05). The gastric mucosa injuries in the rats of rhGH treated group were greatly improved. Conclusion The rhGH, through enhancing the GMBF, can ameliorate the gastric mucosal ischemic-reperfusion injury in rats with hemorrhagic shock.