To study the effects of TNF ? on the protein metabolism and the ubiquitin system gene expression in isolated skeletal muscles, after dissecting and isolating the extensor digitorium longus (EDL) muscles, the in vitro oxygen rich muscle incubation system and as high performance liquid chromatography were used to assess proteolytic rate of the samples. The EDL muscles in study group were incubated with media containing 6 000 U/ml recombinant rat TNF ?. In control group, the media were of the same composition as that of the study group except recombinant rat TNF ?. The expressions of ubiquitin mRNA and C 2 mRNA in rat EDL muscles were determined by Northern blot analysis. No notable difference was observed in the total and myofibrillar proteolytic rate in EDL muscles between the two groups. The expressions of ubiquitin mRNA(2 4kb) and C2 mRNA of EDL muscle incubated with medium containing TNF ? were increased by 151% and 56%, respectively, as compared with those in control group. TNF ? could directly strengthen the function of ubiquitin dependent proteolytic system, but further studies are necessay to elucidate whether TNF ? could directly increase the proteolytic rate in skeletal muscle.

To study the changes of skeletal muscle proteolysis in severely burned patients with sepsis and analyze its possible mechanism, the blood, 24h urine and quadriceps femoris muscle samples were collected from nine severely burned patients with sepsis (the burn sepsis group) and nine patients with plastic operation (the control group). The plasma concentrations of cortisol and TNF ? were determined with radioimmunoassay. The urinary output of 3 methylhistidine (3 MH) in 24h urine were assessed with high performance liquid chromatography. The expressions of ubiquitin mRNA and C 2 mRNA in quadriceps femoris muscle were determined by Northern blot analysis. The protein expressions of ubiquitin in quadriceps femoris muscle were determined by immunopathology. The results showed that the plasma concentrations of cortisol and TNF ? in burn sepsis group were significantly higher than those of the control group ( P

Objective To explore the effect and mechanism of local administration of thymosin β4 (Tβ4) on bone regeneration in a distraction osteogenesis model of rat fenur.Methods Sixty Sprague-Dawley rats were randomly divided into groups A,B and C in this study.A distraction osteogenesis model was established in the left femur after osteotomy.At the end of distraction period,the bone regeneration area in group A was subjected to no treatment,that in group B to injection of phosphate buffer saline (PBS),and that in group C to injection of Tβ4.On days 22,29 and 43 postoperatively,the rats from each group were randomly sacrificed and processed for observation of bone regeneration in the distraction osteogenesis area using radiography,Micro-CT,histology and immunohistochemical staining.RT-PCR was used to detect the expression of related genes as well.Results Radiography revealed that the bone regeneration in group C was superior to that in groups A and B on days 22,29 and 43 postoperatively.Micro-CT examination showed significantly increased bone volume (BV),bone mineral density (BMD) and ratio of bone volume to tissue volume (BV/TV) in group C on days 22,29 and 43 postoperatively,and significantly decreased ratio of bone surface area to bone volume on postoperative day 43 in comparison with groups A and B(P ＜ 0.05).HE staining indicated that local capillary density was significantly higher in group C than in groups A and B after local administration of Tβ4 in the distraction osteogenesis.Immunohistochemical staining showed that the capillary density and osteoblasts in group C were higher than in groups A and B.RT-PCR results revealed significantly higher expression of eNOS and Osterix mRNA in the local callus in group C on postoperative day 22 than in groups A and group B (P ＜ 0.05).Conclusion Local administration of thymosin β4 may promote bone formation,which is probably related to the increased expression of eNOS and Osterix.

Plastic and Aesthetic surgery is a science which creates beauty by combining know-ledge and art. Combined with the professional characteristics of plastic surgery, we reformed the cur-riculum content and teaching mode of continuous education, including a established interactive theoret-ical learning model based on Journal club, the strengthening of clinical practice integrated with multiple related disciplines, the expanding the knowledge of Sociology, Psychology and Ethics, and the construc-tion of a long-term platform of network resources. Therefore, a comprehensive and specialized continu-ous education model in the department of plastic and aesthetic surgery was ultimately formed, whose preliminary assessment was favorable, and could be helpful in the cultivation of high-quality plastic and aesthetic surgeons in the future.

OBJECTIVETo investigate the mechanism and the effects of intravenously injected tumor necrosis factor alpha (TNFalpha) on skeletal muscle protein degradation in rats and its relationship with glucocorticoid.

METHODSForty-five male Wistar rats were randomly divided into 3 groups as A (control), B (TNFalpha injection) and C (TNFalpha and glucocorticoid receptor antagonist injection) groups. TNFalpha in dose of 1x 10(6) units/kg was given to rats in B group intravenously. RU38486, a glucocorticoid receptor antagonist, was given by gavage in C group 2 hours before intravenous injection of TNFalpha in the same dose as in B group. the rat temperature was monitored 12 hours after the administration of the drugs. At the same time, the rat extensor digitorum longus muscles (EDL) were isolated, weighed and cultured under aerobic condition, and than the degradation rates of total and the myofibrillar proteins were determined with HPLC (high performance liquid chromatography), and the expression changes in C2 subunit mRNA and ubiquitin mRNA were detected by Northern blot.

RESULTSTwelve hours after the injection, the temperature of the rats in B and C group was much higher than that in A group (P < 0.01), while the weight of the extensor digitorum longus muscle in B and C groups was evidently lower than that in A group (P < 0.01) whereas that in C was higher than that in B groups (P < 0.05). The degradation rates of total and the myofibrillar proteins in B group were increased by 43% and 112%, respectively, when compared with those in A group (P < 0.01), while the rates in C group was decreased by 16% and 28%, respectively, when compared with those in B group (P < 0.01). In addition, the expressions of ubiquitin mRNA (2.4 kb) and C2 subunit mRNA in B group were increased 4.3 and 3.6 fold compared with those in A group, whereas those in C group were much lower than those in B group.

CONCLUSIONIntravenous injection of recombinant TNFalpha in large dose might enhance the activity of rat skeletal muscle ubiquitin-proteasome system pathway, which led to an increase in the degradation rate of rat total protein, especially the myofibrillar protein. Glucocorticoid was one of the mediating factors of that effect.

Animals ; In Vitro Techniques ; Male ; Muscle Proteins ; metabolism ; Muscle, Skeletal ; drug effects ; Rats ; Rats, Wistar ; Recombinant Proteins ; pharmacology ; Tumor Necrosis Factor-alpha ; pharmacology ; Ubiquitin ; metabolism

Objective To evaluate the role of c-Jun N-terminal kinase (JNK) signaling pathway in paclitaxel-induced apoptosis in hippocampal neurons of rats, and the relationship with nuclear factor kappa B (NF-κB) pathway.Methods The primarily cultured hippocampal neurons were seeded in 96-well plate at a density of 1×106 cells/ml (200 μl/hole) , and were randomly divided into 4 groups (n=8 each) using a random number table: control group (C group), paclitaxel group (P group), JNK inhibitor SP600125 group (S group), and SP600125 + paclitaxel group (S+P group).Paclitaxel 2 ml (1 μmol/L) was added to group P.SP600125 2 ml (10 μmol/L) was added to group S.In group S+P, SP600125 2 ml (10 μmol/L) was added, the cell were then incubated for 1 h, and then paclitaxel 2 ml (1 μmol/L) was added.The cells were then incubated for 24 h.At 24 h of incubation, the apoptosis in hippocampal neurons was detected by flow cytometry, and the expression of NF-κB p65 was measured by Western blot.The apoptosis rate was calculated.Results Compared with group C, the apoptosis rate was significantly increased, and the expression of NF-κB p65 was up-regulated in P and S+P groups, and the apoptosis rate was significantly decreased, and the expression of NF-κB p65 was down-regulated in group S (P＜0.05).Compared with group P, the apoptosis rate was significantly decreased, and the expression of NF-κB p65 was down-regulated in group S+P (P＜0.05).Conclusion JNK signaling pathway mediates paclitaxel-induced apoptosis in hippocampal neurons of rats, and the mechanism is likely related to inhibition of NF-κB pathway activation.

ObjectiveTo investigate the clinical characteristics,epidemiology of patients with severe fever with thrombocytopenia syndrome bunyavirus (SFTSV) infection and genetic sequences of SFTSV.MethodsClinical data of five cases of severe fever with thrombocytopenia syndrome (SFTS)from Zhoushan Hospital during May 2011 to July 2011 were retrospectively analyzed.SFTSV gene was amplified by polymerase chain reaction (PCR).CD3+ CD4+ and CD3+ CD8+T lymphocytes were detected by flow cytometry (FCM).The sequences of isolated SFTSV strains were compared with those in GenBank. ResultsThe symptoms of continuous high fever,sore muscles,enlarged superficial lymph nodes,abdominal pain,diarrhea with gastrointestinal hemorrhage were observed.The white blood cells,platelets and CD3+ CD4+ T lymphocytes were progressive decreased in acute phase with the minimum of (0.97-2.00) × 109/L,(12-42) × 109/L and 7.52％-20.39％,respectively.The SFTSV was isolated from the sera of two patients.The sequences were compared with SFTSV sequences in GenBank.The homology of RNA-dependent RNA polymerase gene was 96％ compared with BX-2010,L-WWG,LN3,JS4,SD4,HN6 and AH12; the glycoprotein gene was 94％ ; N protein gene was 95％ compared with JS4,SD4 and LN4.The homology of the above three genes between two isolates was 99％.ConclusionsOur results suggest that SFTSV is sporadic in Zhejiang Province which is probably from native epidemic focus.SFTS is progressive and severe with acute onset.Multiple organ dysfunction is common in severe eases.

OBJECTIVETo explore the clinical effect of transplantation of the long head of biceps femoris muscle flap in combination with semi-V posterior thigh fasciocutaneous flap for repair of pressure sores over ischial tuberosity.

METHODSEight patients with 10 deep pressure sores over ischial tuberosity were admitted to the First Affiliated Hospital to the PLA General Hospital and the 98th Hospital of PLA from April 2004 to June 2010. The wounds measured from 2 cm × 2 cm to 6 cm × 4 cm were covered with the long head of biceps femoris muscle flap and semi-V posterior thigh fasciocutaneous flap (ranged from 10 cm × 6 cm to 13 cm × 8 cm). The condition of flaps was observed and followed up for a long time.

RESULTSAll flaps survived. Nine wounds healed by first intention. Subcutaneous accumulation of fluids occurred in one wound with formation of a sinus at drainage site, and it healed after dressing change for 25 days. Patients were followed up for 7 to 34 months. Sore recurred in one patient 9 months after surgery, and it was successfully repaired with the same flap for the second time. Flaps in the other 7 patients appeared satisfactory with soft texture and without ulceration.

CONCLUSIONSThis combined flap is easy in formation and transfer, and it causes little side injury with good resistance against pressure. It is a new method for repair of pressure sore over sacral region.

Adult ; Female ; Humans ; Ischium ; Male ; Middle Aged ; Muscle, Skeletal ; transplantation ; Pressure Ulcer ; surgery ; Reconstructive Surgical Procedures ; methods ; Skin Transplantation ; methods ; Surgical Flaps

OBJECTIVETo study the modulatory effect of insulin on apoptosis of skeletal myoblast (L6 cells) by serum of scalded rat and its mechanism.

METHODSL6 cells cultured with DMEM medium containing 10% FBS were divided into control (C, added with 20% normal rat serum), serum from rat with scald injury (S, added with 20% serum from scalded rat), insulin (I, added with 20% normal rat serum and 100 nmol/L insulin), and serum of scalded rat + insulin (SI, added with 20% serum of scalded rat + 100 nmol/L insulin) groups according to the random number table. After being cultured for 48 hours, apoptosis was observed with Hoechst 33258 staining and its number counted, annexin V -FITC/PI double-labeling method was used to assess apoptosis rate, the protein levels of phosphorylated (p-) Akt, p-PI3K, Bax, Bcl-2, and active caspase-3 were determined by Western blotting. Data were processed with grouped or paired t test.

RESULTS(1) The amount of apoptosis with typical morphological change in S group [(59.6 +/- 3.9) per visual field] was more than that in C, I, and SI groups [(4.9 +/- 2.6), (5.5 +/- 2.1), (19.7 +/- 2.3) per visual field, with t value respectively 28.53, 29.86, 21.53, P values all below 0.01]. (2) Apoptotic rate in S group was (18.5 +/- 1.8)%, which was markedly higher than that in C, I, and SI groups [(1.1 +/- 0.6)%, (1.5 +/- 0.3)%, (7.8 +/- 0.6)%, with t value respectively 22.41, 22.83, 13.92, P values all below 0.01]. (3) Compared with those in C group, the protein levels of Bax and active caspase-3 in S group were up-regulated (1.12 +/- 0.63 vs. 0.16 +/- 0.03, 2.15 +/- 0.51 vs. 0.21 +/- 0.03, with t value respectively 3.80, 10.69, P values all below 0.01), the protein level of p-Akt was lowered (0.20 +/- 0.03 vs. 0.42 +/- 0.07, t = -8.46, P < 0.01), and the protein levels of p-PI3K and Bcl-2 showed no statistical difference (0.19 +/- 0.03 vs. 0.26 +/- 0.09, 0.17 +/- 0.03 vs. 0.28 +/- 0.07, with t value respectively -2.73, - 1.14, P values all above 0.05). The protein levels of Bax (0.40 +/- 0.14) and active caspase-3 (0.83 +/- 0.18) in SI group were lowered (t = -3.23, P < 0.05; t = 6.66, P < 0.01) and the protein levels of p-Akt, Bcl-2, and p-PI3K in SI group were elevated (0.39 +/- 0.10, 0.78 +/- 0.03, 0.47 +/- 0.12, with t value respectively 4.07, 18.71, 5.05, P < 0.05 or P < 0.01) as compared with those in S group.

CONCLUSIONSSerum from scalded rat can induce apoptosis in skeletal myoblast, and the effect can be inhibited by insulin through PI3K/Akt signal pathway.

Animals ; Apoptosis ; drug effects ; Burns ; blood ; metabolism ; pathology ; Caspase 3 ; metabolism ; Cell Line ; Insulin ; pharmacology ; Male ; Myoblasts, Skeletal ; cytology ; drug effects ; pathology ; Rats ; Rats, Wistar ; Serum ; immunology ; Signal Transduction ; bcl-2-Associated X Protein ; metabolism

OBJECTIVETo study the regulatory effect of glucagon-like peptide-1 (GLP-1) on cell proliferation of skeletal myoblast strain L6 and its possible signal mechanism.

METHODSL6 cells cultured in DMEM high glucose culture medium containing 10% FBS were divided into control group (C, without addition), GLP-1 group (G, added with 10 nmol/L GLP-1), PI3K inhibitor group (W, added with 50 nmol/L PI3K specific inhibitor wortmannin), and GLP-1 + PI3K inhibitor group (GW, added with 10 nmol/L GLP-1 and 50 nmol/L wortmannin) according to the random number table. Cell proliferation activity was detected with MTT assay at post culture hour (PCH) 24, 48, 72 (denoted as absorbance value). At PCH 24, the change in cell cycle was evaluated with flow cytometer, the expression level of proliferating cell nuclear antigen (PCNA) was determined with immunohistochemical staining, the protein levels of phosphorylated PI3K (p-PI3K) and p-Akt were determined with Western blotting. Data were processed with multi-group analysis of variance.

RESULTS(1) The cell proliferation activity at PCH 48, 72 in G group was respectively 0.660 +/- 0.120, 0.870 +/- 0.240, all significantly higher than those in C group (0.530 +/- 0.060, 0.700 +/- 0.100, with F value respectively 5.46, 5.90, P < 0.05 or P < 0.01). The cell proliferation activity in W group at each time point was lower than that in C group. The cell proliferation activity in GW group at PCH 48, 72 was respectively 0.510 +/- 0.080, 0.740 +/- 0.160, all lower than those in G group (with F value respectively 5.46, 5.90, P < 0.05 or P < 0.01). (2) The percentage of S phase cell in G group at PCH 24 [(15.7 +/- 0.4)%] was significantly higher than that in C group [(13.6 +/- 0.6)%] and GW group [(10.1 +/- 0.6)%], while that in W group [(6.8 +/- 1.2)%] was lower than that in C group (with F values all equal to 15.39, P values all below 0.01). (3) PCNA level in G group at PCH 24 [(51.24 +/- 1.18)%] was markedly higher than that in C group [(36.72 +/- 1.56)%] and GW group [(25.90 +/- 1.22)%], and while in W group [(21.70 +/- 0.09)%] was lower than that in C group (with F values equal to 783.80, P values all below 0.05). (4) The protein level of p-Akt in G group at PCH 24 was significantly higher than that in the other 3 groups, while that in W group was lower than that in C group (with F values equal to 94.43, P values all below 0.01). There was no obvious difference in protein level of p-PI3K at PCH 24 among G, GW, and C groups ( F = 20.94, P > 0.05). The protein level of p-PI3K at PCH 24 in W group was lower than that in C group (F = 20.94, P < 0.05).

CONCLUSIONSGLP-1 can promote cell proliferation of skeletal myoblast by accelerating the progression of cell cycle and increasing the synthesis of DNA, which can be attributed to PI3K/Akt signal pathway.

Androstadienes ; pharmacology ; Animals ; Cell Cycle ; Cell Line ; Cell Proliferation ; drug effects ; DNA ; biosynthesis ; Glucagon-Like Peptide 1 ; pharmacology ; Myoblasts, Skeletal ; drug effects ; metabolism ; pathology ; Phosphatidylinositol 3-Kinases ; antagonists & inhibitors ; metabolism ; Rats ; Signal Transduction ; drug effects