Effects of human ß-defensin-3 on biofilm formation­regulating genes dltB and icaA in Staphylococcus aureus.

An understanding of the regulatory mechanisms that drive Staphylococcus aureus biofilm formation may lead to the development of an effective strategy to control the increasing number of refractory clinical infections it causes. The present study examined the effects of the antimicrobial agent human ß­defensin 3 (hBD­3) and the antibiotics vancomycin and clindamycin on the expression of the S. aureus biofilm formation­regulating genes, icaA and dltB, during bacterial adhesion and biofilm formation. Transcription (mRNA) levels of dlt and ica genes were measured using quantitative polymerase chain reaction, and slimes of S. aureus biofilm were examined with confocal scanning laser microscopy during S. aureus adhesion and biofilm formation. Although hBD­3, vancomycin and clindamycin led to significantly attenuated biofilm formation, their treatment­associated effects on the mRNA expression of dlt and ica were not identical. Vancomycin and clindamycin induced sustained expression of the dlt and ica genes, which may be harnessed to induce biofilm formation. However, hBD­3 did not have a significant affect on the transcription level of dltB during either bacterial adhesion or biofilm formation. Therefore, the mechanism of hBD­3 that regulated the suppression of biofilm formation appears to differ from the mechanisms of vancomycin and clindamycin.

Human beta-defensin-3 for the diagnosis of periprosthetic joint infection and loosening.

In this study, the difference in expression of human beta-defensin-3 in periprosthetic tissue and cancellous bone was observed in the periprosthetic tissue and cancellous bone of patients in the periprosthetic joint infection group, the aseptic loosening group, and the spacer treatment group as well as the synovial membrane and ilium of the normal control group. Hematoxylin and eosin staining of the synovial tissue showed different levels of neutrophil infiltration in all groups except for the normal control group. Immunofluorescence staining of periprosthetic tissue and cancellous bone showed the most positive cells expressing human beta-defensin-3 and the largest mean optical density in the periprosthetic joint infection group, followed by the aseptic loosening group, the spacer treatment group, and the normal control group, with a significant difference in comparison between the periprosthetic joint infection group and the other 3 groups (P<.01). White blood cell count, erythrocyte sedimentation rate, and C-reactive protein level were highest in the periprosthetic joint infection group, whereas no difference was found between the other 3 groups. Taken together, high levels of human beta-defensin-3 protein expression were found in the periprosthetic tissue and cancellous bone of patients with periprosthetic joint infection and aseptic loosening, but there are differential expressions of human beta-defensin-3, and this may provide a new marker for the differential diagnosis of infection and loosening of the artificial joint.

Peripheral leukocytapheresis attenuates acute lung injury induced by lipopolysaccharide in vivo.

The mortality of acute lung injury and acute respiratory distress syndrome (ALI/ARDS) remains high and efforts for prevention and treatments have shown little improvement over the past decades. The present study investigated the efficacy and mechanism of leukocytapheresis (LCAP) to partially eliminate peripheral neutrophils and attenuate lipopolysaccharide (LPS)-induced lung injury in dogs. A total of 24 healthy male mongrel dogs were enrolled and randomly divided into LPS, LCAP and LCAP-sham groups. All animals were injected with LPS to induce endotoxemia. The serum levels of leucocytes, neutrophil elastase, arterial blood gas, nuclear factor-kappa B (NF-κB) subunit p65 in lung tissues were measured. The histopathology and parenchyma apoptosis of lung tissues were examined. We found that 7, 3, and 7 animals in the LPS, LCAP, and sham-LCAP groups, respectively, developed ALI 36 h after LPS infusion. The levels of NF-κB p65 in lung tissue, neutrophils and elastase in blood, decreased significantly following LCAP. LCAP also alleviated apoptosis, and NF-κB p65 in lung tissues. Collectively, our results show that partial removal of leucocytes from peripheral blood decreases elastase level in serum. This, in turn, attenuates lung injuries and may potentially decrease the incidence of ALI.

Comparison of the effects of human ß-defensin 3, vancomycin, and clindamycin on Staphylococcus aureus biofilm formation.

Despite improvements in surgical techniques and implant design in orthopedic surgery, implantation-associated infections are still a challenging problem for surgeons. In 2006, trace quantities of human ß-defensin 3 (hBD-3) were found in human bone tissue and bone cells. Human ß-defensin 3 is a 45-amino-acid peptide that is considered the most promising class of defensin antimicrobial peptides and may help in the prevention and treatment of implantation-associated infections. Studies of the effectiveness of hBD-3 against Staphylococcus aureus showed that hBD-3 was more potent at low concentrations than other antibiotics. The effect of hBD-3 on S aureus biofilms has not been reported. We studied the effect of hBD-3, vancomycin, and clindamycin on S aureus biofilms and on the survival of the bacteria in the biofilms.Staphylococcus aureus biofilms were examined with confocal scanning laser microscopy. Staining with LIVE/DEAD BacLight viability stain (Molecular Probes Europe BV, Leiden, The Netherlands) differentiated between live and dead bacteria within the biofilms, and extracellular polymeric substances (slime) from the biofilms was evaluated after staining with calcofluor white (Sigma Chemical Company, Rocky Hill, New Jersey). Human ß-defensin 3 and clindamycin reduced the S aureus biofilm area. Human ß-defensin 3 was significantly more effective against bacteria from the S aureus biofilms than was clindamycin. Vancomycin did not reduce the S aureus biofilm area.

Surgical treatment strategy for multiple injury patients in ICU.

OBJECTIVE: To investigate the surgical treatment for patients with multiple injuries in ICU. METHODS: Clinical data of 163 multiple injury patients admitted to ICU of our hospital from January 2006 to January 2009 were retrospectively studied, including 118 males and 45 females, with the mean age of 36.2 years (range, 5-67 years). The injury regions included head and neck (29 cases), face (32 cases), chest (89 cases), abdomen (77 cases), pelvis and limbs (91 cases) and body surface (83 cases). There were 57 cases combined with shock. ISS values varied from 10 to 54, 18.42 on average. Patients received surgical treatments in ICU within respectively 24 hours (10 cases), 24-48 hours (8 cases), 3-7 days (7 cases) and 8-14 days (23 cases). RESULTS: For the 163 patients, the duration of ICU stay ranged from 2 to 29 days, with the average value of 7.56 days. Among them, 143 were cured (87.73%), 11 died in the hospital (6.75%) due to severe hemorrhagic shock (6 cases), craniocerebral injury (3 cases) and multiple organ failure (2 cases), and 9 died after voluntarily discharging from hospital (5.52%). The total mortality rate was 12.27%. CONCLUSIONS: The damage control principle should be followed when multiple injury patients are resuscitated in ICU. Surgical treatment strategies include actively controlling hemorrhage, treating the previously missed injuries and related wounds or surgical complications and performing planned staging operations.

Relationship between disseminated intravascular coagulation and levels of plasma thrombinogen segment 1+2, D-dimer, and thrombomodulin in patients with multiple injuries.

OBJECTIVE: To explore the relationship between disseminated intravascular coagulation (DIC) and levels of plasma thrombinogen segment 1+2 (F1+2), D-dimer (D-D), and thrombomodulin (TM) in patients with severe multiple injuries. METHODS: In this study, 66 patients (49 males and 17 females, aged 15-74 years, mean=38.4 years) with multiple injuries, who were admitted to our hospital within 24 hours after injury with no personal or family history of hematopathy or coagulopathy, were divided into a minor injury group (ISS<16, n=21) and a major injury group (ISS>or=16, n=45) according to the injury severity. The patients in the major injury group were divided into a subgroup complicated with DIC (DIC subgroup, n=12) and a subgroup complicated with no DIC (non-DIC subgroup, n=33). Ten healthy people (7 males and 3 females, aged 22-61 years, mean=36.5 years+/-9.0 years), who received somatoscopy and diagnosed as healthy, served as the control group. Venous blood samples were collected once in the control group and 1, 3 and 7 days after trauma in the injury groups. The F1+2 and TM concentrations were determined by enzyme linked immunosorbent assay (ELISA), and D-D concentrations were measured by automated latex enhanced immunoassay. RESULTS: F1+2, D-D and TM levels were higher in the minor and major injury groups than in the control group. They were markedly higher in the major injury group than in the minor injury group. In the non-DIC subgroup, F1+2 levels declined gradually while D-D and TM levels declined continuously. In the DIC subgroup, F1+2 and D-D levels remained elevated while TM levels exhibited an early rise and subsequent decrease. Plasma F1+2, D-D and TM levels were higher in the DIC patients than in the non-DIC patients. Injury-induced increases in F1+2, D-D and TM plasma levels had significant positive correlation with each other at each time point. CONCLUSIONS: Besides being related to trauma severity, F1+2, D-D and TM levels correlate closely with the occurrence of posttraumatic DIC. Therefore, changes in plasma F1+2, D-D and TM levels may predict the occurrence of DIC.

Damage control surgery for severe thoracic and abdominal injuries / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To investigate the application of damage control surgery in treatment of patients with severe thoracic and abdominal injuries.</p><p><b>METHODS</b>A retrospective study was done on 37 patients with severe thoracic and abdominal injuries who underwent damage control surgery from January 2000 to October 2006 in our department. There were 8 cases of polytrauma (with thoracic injury most commonly seen), 21 of polytrauma (with abdominal injury most commonly seen) and 8 of single abdominal trauma. Main organ damage included smashed hepatic injuries in 17 cases, posterior hepatic veins injuries in 8,pancreaticoduodenal injuries in 7, epidural or subdural hemorrhage in 4, contusion and laceration of brain in 5, severe lung and bronchus injuries in 4, pelvis and one smashed lower limb wound in 3 and pelvic fractures and retroperitoneal hemorrhage in 6. Injury severity score (ISS) was 28-45 scores (38.4 scores on average), abbreviated injury scale (AIS) > or = 4.13. The patients underwent arteriography and arterial embolization including arteria hepatica embolization in 4 patients, arteria renalis embolization in 2 and pelvic arteria retroperitoneal embolization in 7. Once abbreviated operation finished, the patients were sent to ICU for resuscitation. Twenty-four cases underwent definitive operation within 48 hours after initial operation, 5 underwent definitive operation within 72 hours after initial operation, 2 cases underwent definitive operation postponed to 96 hours after initial operation for secondary operation to control bleeding because of abdominal cavity hemorrhea. Two cases underwent urgent laparotomy and decompression because of abdominal compartment syndrome and 2 cases underwent secondary operation because of intestinal fistulae (1 case of small intestinal fistula and 1 colon fistula) and gangrene of gallbladder.</p><p><b>RESULTS</b>A total of 28 patients survived, with a survival rate of 75.68%, and 9 died (4 died within 24 hours and 5 died 3-9 days after injury). The trauma deaths at the early stage were caused by severe primary injuries resulting in failure of respiration and circulation, while mortality at the later stage was caused by multiple organ failure.</p><p><b>CONCLUSIONS</b>Damage control surgery is important for the treatment against severe thoracic and abdominal injuries. It is suggested that the surgeon should select the reasonable auxiliary examination before operation, and take the proper time to perform damage control and definitive surgery.</p>

[Effects of glutamine granules on immunofunction in trauma patients: a double-blind randomized controlled, multi-center clinical trail with 120 patients].

OBJECTIVE: To evaluate the effect of glutamine granules on immunofunction in severe burns and trauma patients. METHODS: One hundred and twenty patients with severe burns, multiple trauma and post operation who met the requirements of the protocol joined this double-blind randomized controlled, multi-center clinical trail. Patients were randomly divided into two groups: placebo control group (P group, 60 patients) and glutamine granules treatment group (GLN group, 60 patients). There was isonitrogenous and isocaloric intake in both groups. GLN and P group patients had been given glutamine granules or placebo (glycine) at 0.5 g.kg(-1).d(-1) for 7 days, respectively. The level of plasma glutamine and some index of immunofunction were determined, and the complication and side effect were also observed. RESULTS: After 7 days of taking glutamine granules orally, plasma GLN concentration was significantly higher than that in P group [(593 +/- 185) micromol/L vs (407 +/- 190) micromol/L)] (P < 0.01). IL-2 level, CD(4)/CD(8) ratio, PMN swallow ratio in GLN group were significantly higher than those in P group (P < 0.05-0.01), but the concentration of IgG, IgM, C(3)/C(4) were not significantly different when compared with P group (P > 0.05). In addition, the occurrence of side effect in both groups was seldom. CONCLUSION: Taking glutamine granules could increase plasma GLN concentration, enhance body immunofunction, and using glutamine granules is safe.

[Study on high mobility group-1 protein in patients with multiple trauma].

OBJECTIVE: To study the change in serum high mobility group-1 (HMG-1) protein in patients with multiple trauma and determine its relationship to severity of trauma and multiple organ dysfunction syndrome (MODS). METHODS: In 33 cases, HMG-1 levels were determined on 1, 3, 7 days after trauma respectively. The values of injury severity score (ISS), acute physiology and chronic health evaluation II (APACHE II) and systemic inflammatory response syndrome (SIRS) score were determined. Meanwhile, the occurrence of organ dysfunction was noted. RESULTS: The level of serum HMG-1 in patients with multiple trauma was much higher than that of normal controls, and also in those with organ dysfunction. HMG-1 and APACHE II were positively correlated (r=0.495, P=0.016). The level of serum HMG-1 and APACHE II value were much higher in patients with organ dysfunction than those of normal organ function, but there was no significant difference in SIRS score between the two groups (P=0.105). CONCLUSION: Serum HMG-1 increases in patients with trauma and is positively correlated with severity of trauma. Serial determination of HMG-1 is helpful to discover clinical infection earlier. HMG-1 can be used as a warning indicator of the onset of MODS.

[Analysis of the therapeutic effect and the safety of glutamine granules per os in patients with severe burns and trauma].

OBJECTIVE: To observe the therapeutic effect and possible side effects of glutamine granules per os in patients with trauma, burns and major operations. METHODS: Patients inflicted with severe burns, trauma and major operations were enrolled in the study. One hundred and twenty patients were randomly divided into two groups, 60 in control group (C) and 60 in glutamine group (Gln). Randomized double blind and placebo control methods were employed in the study. All the patients in both groups were given diet with equal calories and equal nitrogen content. The patients in Gln group received glutamine granules in dose of 0.5 g.kg(-1).d(-1) orally or by gavage, while those in C group received same dose of placebo (glycine) for 7 days. The changes in the intestinal mucosal barrier function, the protein metabolism, the immune function, hepatic and renal functions, and the incidence of side effects of the medication in both groups of patients were observed and compared before and after the supplementation of glutamine or glycine. RESULTS: The plasma contents of glutamine, proteins and interleukin 2 in both groups were all lower than normal values. But the plasma diamine oxidase (DAO) activity, endotoxin content, intestinal mucosal permeability (urine lactose/mannitol, L/M) and urine excretion of nitrogen increased obviously in both groups. The plasma glutamine concentration in Gln group increased by 38.04% after the administration of Gln for 7 days (P < 0.01). The plasma contents of pro-albumin, transferrin, and IL-2 were obviously higher than those in the C group (the increase rates were 21.19%, 51.11%, 57.54%, respectively, P < 0.01). The plasma DAO activity, L/M ratio, endotoxin content and urine nitrogen excretion in Gln group were evidently lower than those in C group (the decrease rates were 47.26%, 52.18, 22.22% and 27.78%, respectively, P < 0.05 or 0.01). There was no obvious difference in the plasma levels of total protein and albumin, the indices in blood and urine test, or the hepatic and renal functions between the two groups before and after the amino acid supplementation. Mild side effects such as nausea, diarrhea, constipation occurred in both groups, but all of them disappeared spontaneously afterwards (P > 0.05). CONCLUSION: Oral administration of glutamine could be helpful to increase plasma concentration of glutamine and to ameliorate obviously the intestinal mucosal injury, to promote systemic protein synthesis and to inhibit protein catabolism and to upgrade systemic immune function with little side effect in patients with severe injury.

[Effects of glutamine granules on protein metabolism in trauma patients].

OBJECTIVE: To evaluate the effect of glutamine granules on protein metabolism in severe burns and trauma patients. METHODS: 120 patients with severe burns, multiple trauma and post operation who met the requirements of the protocol joined this double-blind randomized controlled, multi-center clinical trail. Patients were randomly divided into two groups: placebo control group (P group, 60 patients) and glutamine granules treatment group (GLN group, 60 patients). There was isonitrogenous and isocaloric intake in both groups, GLN and P group patents had been given glutamine granules or placebo (glycine) at 0.5 g.kg(-1).d(-1) for 7 days, respectively. The level of plasma glutamine, protein and urine nitrogen exclude were determined, wound healing rate of burn area and hospital stay were recorded, and then observed the complication and side effect. RESULTS: After 7 days of taking glutamine granules orally, plasma GLN concentration was significant higher than that in P group (592.50 +/- 185.23 micro mol/L vs. 407.41 +/- 190.22 micro mol/L) (P < 0.01). Plasma prealbumin and transferrin in GLN group were significant higher than those in P group (P < 0.01), but the concentration of total protein and albumin were no marked changes compare with P group (P > 0.05). The capacity of urine nitrogen exclude in GLN group were significant lower than that in P group. Additional, the wound healing rate was faster and hospital stay days was shorter than P group (P < 0.05), and the occurrence of glutamine granules side effect was seldom. CONCLUSION: Taking glutamine could promote protein synthesis, abate protein decompose, ameliorate wound healing rate and reduce hospital stay obviously.

[Treatment of open fracture by vacuum sealing technique and internal fixation].

OBJECTIVE: To investigate the effect of vacuum sealing(VS) technique and emergency internal fixation on the management of limbs open fracture and soft tissue dirty defects. METHODS: Fourteen patients (18 limbs) with open fracture and soft tissue dirty defects were treated by the VS technique and internal fixation after debridement and 14 patients managed by traditional method as control group. Wound surface were covered with polyvinyl alcohol foams with embedded drainage tubes connected with vacuum bottle (negative pressure of 50 to 60 kPa) after wound surface were debrided and fracture were fixed. Wound closure was performed with secondary suturing, or free flap, or loco-regional flap and mesh-grafts after 5 to 7 days. RESULTS: All wound surface healed completely. No complications (systemic and local) were found. After 4-6 months follow-up on average, the fracture healed well. There was significant difference in time of treatment, total cost of treatment and complication rate between 2 groups (P < 0.01). CONCLUSION: The VS procedure can drain the wound surface completely, decrease infection rate and stimulate the proliferation of granulation tissue. A combination of VS with emergency internal fixation is a simple and effective method in treatment of limbs open fracture and soft tissue dirty defects.

[Study on the relationship of heat shock protein 70 with secondary liver damage in early stage after severe multiple injury].

OBJECTIVE: To study the action of heat shock protein 70 (HSP70) in the course of secondary hepatic injury in rats in the early stage after multiple injury. METHODS: Adult male Wistar rats were used and rat model was produced by adopting severe thoracic impact injury in accompany with mono femur fracture. Changes of the content of HSP 70 in the hepatic tissue, hepatic function markers in serum were dynamically observed as well as 24 hours mortal rate after severe multiple injury, trauma with glucocorticoid receptor(GR) blocked. The expression of HSP70 in hepatic tissue was assayed by western blot and then analyzed with computer imaging system. RESULTS: The content of HSP70 gradually increased in hepatic tissue after severe trauma, increased to the highest at 8 hours after trauma, and maintained rather high level at 24 hours after trauma. But the alterations of hepatic function markers in serum were not obvious after trauma. After usage of GR blocking agent, The content of HSP70 was much higher than simple trauma group. Alanine aminotransferase (ALT) and total bilirubin (TB) in serum obviously increased in early stage after trauma (both P<0.01), albumin content obviously decreased(P<0.01), while 24 hours mortal rate obviously increased (P<0.01). CONCLUSION: HSP70 may participate in starting a anti-injury mechanism of hepatic tissue cell. But excessive expression of HSP70 may cause injury to liver after severe multiple injury. HSP70 can be identified as an important marker of liver injury and anti-injury after severe multiple injury.

[Treatment of traumatic soft tissue defect by vacuum sealing].

OBJECTIVE: To investigate the clinical effect of vacuum sealing in treatment of traumatic soft tissue defect. METHODS: From 1998. 8 to 2001. 2, 49 patients with 55 traumatic soft tissue defects were treated by vacuum sealing after debridement. Among them, there were 39 males and 10 females with mean age 38. 4 years. The wound area varied from 10 cm x 10 cm to 30 cm x 30 cm. In the experimental group, the wound surfaces or cavities were filled with polyvinyl alcohol foams with embedded drainage tubes connected with vacuum bottle (negative pressure of 50-60 kPa). Wound closure was performed with secondary suturing, or skin transplantation, or local flap grafting after 5-7 days. Besides, 126 patients were managed by traditional dressing as the control group. RESULTS: Out of 51 traumatic soft tissue defects (45 patients), the wound closure was performed with a free flap in 4, with local flap in 8, with skin grafting in 27, with secondary suturing in 6, and with vacuum sealing directly in the other 6 defects. Forty-five patients recovered and no complications (systemic and located) occurred. There were significant differences in time of secondary suturing, times of dressing, wound shrink and total therapeutic cost between two groups (P < 0.01). CONCLUSION: Vacuum sealing can protect the wound against contamination, evacuate the wound exudates completely, stimulate the growth of granulation tissue, and facilitate the wound healing; so vacuum sealing is a simple and effective method in treatment of traumatic soft tissue defect.