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Object. This study was performed to determine whether moderate hypothermia (31°C) improves clinical outcome in severely head injured patients whose intracranial hypertension cannot be controlled using mild hypothermia (34°C). Methods. Twenty-two consecutive severely head injured patients who fulfilled the following criteria were included in this study: an intracranial pressure (ICP) that remained higher than 40 mm Hg despite the use of mild hypothermia combined with conventional therapies; and a Glasgow Coma Scale score of 8 or less on admission. After the failure of mild hypothermia in combination with conventional therapies; patients were exposed to moderate hypothermia as quickly as possible. As brain temperature was reduced from 34 to 31°C, the volume of intravenous fluid infusion was increased significantly from 1.9 ± 0.9 to 2.6 ± 1.2 mg/kg/hr (p < 0.01), and the dose of dopamine infusion increased significantly from 4.3 ± 3.1 to 8.2 ± 4.4 µg/kg/min (p < 0.01). Nevertheless, mean arterial blood pressure and heart rate decreased significantly from 97.1 ± 13.1 to 85.1 ± 10.5 mm Hg (p < 0.01) and from 92.2 ± 13.8 to 72.2 ± 14.3 beats/minute at (p < 0.01) at 34 and 31°C, respectively. Arterial base excess was significantly aggravated from −3.3 ± 4 at 34°C to −5.6 ± 5.4 mEq/L (at 31°C; p < 0.05). Likewise, serum potassium concentration, white blood cell counts, and platelet counts at 31°C decreased significantly compared with those at 34°C (p < 0.01). In 19 (86%) of 22 patients, elevation of ICP could not be prevented using moderate hypothermia. In the remaining three patients, ICP was maintained below 40 mm Hg by inducing moderate hypothermia; however, these three patients died of multiple organ failure. These results clearly indicate that moderate hypothermia induces complications more severe than those induced by mild hypothermia without improving outcomes. Conclusions. The authors concluded that moderate hypothermia is not effective in improving clinical outcomes in severely head injured patients whose ICP remains higher than 40 mm Hg after treatment with mild hypothermia combined with conventional therapies.

Introduction The gut is an important target organ after severe insult. Gut microbiota have an important role in immune response. However, the gut microbiota and environment have not been clarified in patients with burns. Therefore, we serially evaluated the gut microbiota and environment in patients with major burns. Methods Fecal samples from five patients with major burns were measured for quantitative evaluation of the gut microbiota. Results In the four survivors of major burns, the numbers of beneficial bacteria, especially those of total obligate anaerobes and Bifidobacterium, initially decreased, but then increased as the condition of the survivors improved. By contrast, the numbers severely decreased in the non-survivor as gut failure and sepsis progressed. The number of pathogenic bacteria such as Pseudomonas and Candida did not continue to increase in the survivors, whereas in the non-survivor the number increased and continued to higher counts. Short-chain fatty acids such as propionic and butyric acids decreased to lower-than-normal levels but tended to increase after recovery in the survivors. The levels remained below normal in the non-survivor. Conclusions The gut microbiota and environment are severely altered in patients with major burns. Consequently, abnormal gut conditions may have an influence on the systemic inflammatory response and multiple organ dysfunction syndrome. A novel treatment to maintain the gut microbiota and environment is expected in the future.

Aquaporins (AQPs) are water channels of cell membranes. All living cells experience osmotic pressure changes in their environment, but the mechanism by which water influx occurs was not known until the discovery of AQPs. AQP9, which is expressed in human polymorphonuclear leukocytes (PMNLs), is reported to relate to morphologic changes of PMNLs in vitro. We examined the expression of AQP9 in PMNLs from patients with systemic inflammatory response syndrome (SIRS) and addressed the role of AQP9 in both morphologic and functional changes of PMNLs in the SIRS condition. Fourteen patients with SIRS were included in our study. Polyclonal antibody was used for the AQP9 assay. F-actin polymerization, oxidative activity, and the expression of AQP9 in PMNLs with and without stimulation by N-formylmethionyl-leucyl-phenylalanine were evaluated by flow cytometry. Expression of AQP9, F-actin polymerization, and oxidative activity in PMNLs were increased significantly in patients with SIRS compared with those in healthy volunteers. The time course of AQP9 fluorescence in PMNLs corresponded to the time course of F-actin polymerization, which showed peak fluorescence at 1 min after N-formylmethionyl-leucyl-phenylalanine stimulation. The expression of AQP9 in PMNLs is increased significantly in SIRS patients. The increased expression of AQP9 in SIRS patients might be associated with F-actin polymerization in PMNLs, which could affect both morphologic and functional changes of PMNLs in the SIRS condition.

肝細胞癌の術中に偶然発見した最大径1cmの細小胆管細胞癌の1例を経験したので報告する.症例は64歳,男性. C型肝硬変の経過観察中に3個の肝細胞癌を指摘されて,当科に紹介された. S4, S5, S7の肝細胞癌の診断で手術を施行した. S4, S7の部分切除術, S5区域切除術を予定していたが, S5区域切除時に肝切離面に白色調の硬結を認めたため,切離線を変更し同病変も併せて切除した.病理組織学的に術前診断した3病変はいずれも中分化型肝細胞癌,術中に発見した病変は中分化型管状腺癌であり,腫瘤形成型の細小胆管細胞癌と診断した.さらにこの中分化型管状腺癌は神経およびリンパ管に浸潤していた.本症例は肝細胞癌と胆管細胞癌の重複癌であり,胆管細胞癌は1.0cm大と小さいにも関わらず進行癌としての所見を呈しており,興味ある症例と考えられた.

In comprehensive glycome analysis with a high-density lectin microarray, we have previously shown that the recombinant N-terminal domain of the lectin BC2L-C from Burkholderia cenocepacia (rBC2LCN) binds exclusively to undifferentiated human induced pluripotent stem (iPS) cells and embryonic stem (ES) cells but not to differentiated somatic cells. Here we demonstrate that podocalyxin, a heavily glycosylated type 1 transmembrane protein, is a glycoprotein ligand of rBC2LCN on human iPS cells and ES cells. When analyzed by DNA microarray, podocalyxin was found to be highly expressed in both iPS cells and ES cells. Western and lectin blotting revealed that rBC2LCN binds to podocalyxin with a high molecular weight of more than 240 kDa in undifferentiated iPS cells of six different origins and four ES cell lines, but no binding was observed in either differentiated mouse feeder cells or somatic cells. The specific binding of rBC2LCN to podocalyxin prepared from a large set of iPS cells (138 types) and ES cells (15 types) was also confirmed using a high-throughput antibody-overlay lectin microarray. Alkaline digestion greatly reduced the binding of rBC2LCN to podocalyxin, indicating that the major glycan ligands of rBC2LCN are presented on O-glycans. Furthermore, rBC2LCN was found to exhibit significant affinity to a branched O-glycan comprising an H type 3 structure (Ka, 2.5 × 104 M−1) prepared from human 201B7 iPS cells, indicating that H type 3 is a most probable potential pluripotency marker. We conclude that podocalyxin is a glycoprotein ligand of rBC2LCN on human iPS cells and ES cells.

Objective: A real-time polymerase chain reaction (PCR) test is expected for early and precise detection of pathogens in blood. In this study, we compared the ability of the PCR test and blood culture to detect pathogens in the blood of patients with sepsis. Methods: Patients who were diagnosed as or suspected of having sepsis were included in this prospective observational study. A whole blood sample for PCR test was obtained serially simultaneously with the blood culture sample, and the results were compared. Results: We obtained 93 samples from 26 patients; 69 samples were obtained during the septic condition, and 24 samples were from the non-septic condition. Origins of sepsis were pneumonia in 9 patients, necrotizing fasciitis in 5 patients, and other causes in 12 patients. In the septic condition, rates of positive results were 29.0% for the PCR test and 23.2% for blood culture. Sample contamination occurred in 1 PCR test sample and 5 blood culture samples. In positive PCR samples, 16 of 20 samples were obtained during sepsis after beginning administration of broad- spectrum antibiotics, whereas 5 of 12 samples, except for contaminated samples, were positive in blood culture. Conclusion: In sepsis, the PCR test detected more bacteria than did blood culture even after administration of empirical antibiotics, which might contribute to precise diagnosis of the bacteremic cause of sepsis

Induced pluripotent stem cells (iPSCs) can now be produced from various somatic cell (SC) lines by ectopic expression of the four transcription factors. Although the procedure has been demonstrated to induce global change in gene and microRNA expressions and even epigenetic modification, it remains largely unknown how this transcription factor-induced reprogramming affects the total glycan repertoire expressed on the cells. Here we performed a comprehensive glycan analysis using 114 types of human iPSCs generated from five different SCs and compared their glycomes with those of human embryonic stem cells (ESCs; nine cell types) using a high density lectin microarray. In unsupervised cluster analysis of the results obtained by lectin microarray, both undifferentiated iPSCs and ESCs were clustered as one large group. However, they were clearly separated from the group of differentiated SCs, whereas all of the four SCs had apparently distinct glycome profiles from one another, demonstrating that SCs with originally distinct glycan profiles have acquired those similar to ESCs upon induction of pluripotency. Thirty-eight lectins discriminating between SCs and iPSCs/ESCs were statistically selected, and characteristic features of the pluripotent state were then obtained at the level of the cellular glycome. The expression profiles of relevant glycosyltransferase genes agreed well with the results obtained by lectin microarray. Among the 38 lectins, rBC2LCN was found to detect only undifferentiated iPSCs/ESCs and not differentiated SCs. Hence, the high density lectin microarray has proved to be valid for not only comprehensive analysis of glycans but also diagnosis of stem cells under the concept of the cellular glycome.

We have developed an expanded polytetrafluoroethylene polymer (e-P) coated with both poly-amino-acid urethane copolymer and collagen (e-PPC) to be used for cell culture substrata. Rat mesenchymal stem cells (MSC) were cultured on the e-P and e-PPC polymers in the presence of dexamethasone. After 24 h, the MSC contacted well with the e-PPC surface and after 14 days, the MSC showed high levels of alkaline phosphatase activity, and calcium and bone-specific osteocalcin protein deposition. The MSC cultured on these polymers for 1 week were then implanted at rat subcutaneous sites and harvested after 4 weeks. Microcomputed tomography as well as histological analyses showed that any hard tissue could not be seen in implants of the MSC/e-P composites, whereas new bone formation could be detected in the MSC/e-PPC composites. These in vitro as well as in vivo results confirmed the importance of polymer surface to support the osteogenic differentiation, which resulted in new bone formation. The surface modification using poly-amino-acid urethane copolymer and collagen together with tissue engineering technology might facilitate bone anchoring to the polymers for dental and orthopedic applications.

Background: The mechanism of blood flow during chest compression in cardiac arrest patients remains under investigation. We often experience that cardiac arrest patients suffer severe diarrhea after successful cardiopulmonary resuscitation(CPR), which would be attributable to intestinal ischaemia during cardiac arrest and resuscitation. However, few studies have been made to evaluate abdominal blood flow during chest compression in cardiac arrest patients. Patients and Methods: The study was made in four patients immediately after termination of cardiopulmonary resuscitation. A 100ml bolus of 300 mgI/ml contrast medium was injected from a short femoral vein catheter, followed by continuous chest compression at a rate 100/min. In order to evaluate the distribution of contrast medium, CT images of the chest and abdomen were taken after 100 and/or 200 chest compressions, respectively. Results: CT scans showed similar enhance patterns in the patients. After 100 chest compressions, enhancement values were higher at inferior vena cava(IVC) regions compared to ascending aorta and main arteries. After 200 compressions, enhancement values of the arteries were increased by two to four times. However, significant enhancement of hepatic veins and limited enhancement of portal veins suggest impaired perfusion of the liver and the bowel. Conclusion: One hundred or 200 chest consecutive compressions are not sufficient to enhance abdominal organs including the liver and bowel. Figures: Typical MPR(Multiplanar reconstruction) images after 100compressions at two different coronal planes; hepatic vein and IVC(left), portal veins(right) are depicted.

Ventilator-associated pneumonia (VAP) is a common and serious problem in ICUs. Several studies have been conducted to determine the effectiveness of Gram stain of tracheal aspirates for diagnosing VAP. However, the effectiveness for predicting causative microorganisms and guiding appropriate initial antibiotic therapy has not been evaluated. The purpose of this study is to determine whether Gram stain of tracheal aspirates can guide appropriate initial antibiotic therapy for VAP.

Smoke inhalation is a significant comorbid factor following major thermal injury. Smoke exposure is only a trigger for the sequence of events responsible for the development of inhalation injury. Noxious chemicals generated by incomplete combustion injure the exposed bronchoepithelium and stimulate the release of chemical mediators that cause a progressive inflammatory process. Airway inflammation and pulmonary edema impair gas exchange and increase the susceptibility to pulmonary infection. Earlier diagnosis and treatment of inhalation injury is an important element to improve the clinical course of severe burn patients. The American Burn Association, however, recently concluded that there are insufficient data to support a treatment standard for the diagnosis of inhalation injury. At present, the diagnosis of inhalation injury is supported by the combination of history, physical examination, bronchoscopy, and laboratory findings For accurate diagnosis of inhalation injury, helical CT scanning and examination to detect activated leukocytes in bronchoalveolar lavage fluid may be warranted. In the respiratory management of inhalation injury, repeated removal of pseudomembrane by fiberoptic bronchoscopy and the use of adequate PEEP to avoid airway obstruction are essential. High-frequency percussive ventilation can be a suitable mode of ventilation for inhalation injury.

The in vitro and in vivo osteoblastic differentiation of rat bone marrow stromal cells (MSCs) was assessed on hydroxyapatite disks with 3 different porosities: 30%, 50%, and 70% (HA30, HA50, and HA70, respectively). MSCs obtained by 10-day culture of fresh bone marrow cells were subcultured for 2 weeks on 3 kinds of porous HA disks in the presence and absence of dexamethasone (Dex). After 2 weeks of subculture, alkaline phosphatase (ALP) activity and osteocalcin production of MSCs/HA composites with Dex were higher than those without, and increased with increasing porosity. The resultant bone tissue grafts “cultured-bone/HA constructs” were implanted subcutaneously into the backs of syngeneic rats, and harvested 1, 2, and 4 weeks after implantation. At 1 week, only cultured-bone/HA70 constructs exhibited expanded bone formation. At 2 and 4 weeks, active osteoblasts and progressive bone formation were observed morphologically in both cultured-bone/HA50 and HA70 constructs. At 4 weeks, bone tissue was observed even in cultured-bone/HA30 constructs. ALP activity and osteocalcin production also increased with increasing porosity and time after implantation. In this in vivo model, different scaffold porosity with similar crystal morphology of the apatite phase demonstrated marked differences in ability to support osteogenesis by implanted rat MSCs.

Innate immunity plays an important role in host defense after severe insult. gammadelta T lymphocytes are recognized as the first line of defense against microbial invasion. In this study, we evaluated gammadelta T lymphocytes in the peripheral blood of patients with severe systemic inflammatory response syndrome (SIRS), and examined on role of these cells. Thirty-seven patients with severe SIRS (SIRS criteria and serum C-reactive proteinor = 10 mg/dL) and 27 healthy volunteers were studied. Severe SIRS was caused by trauma in 14 patients (Injury Severity Score of 30.1 +/- 10.8) and by sepsis in 23 patients. The counts of gammadelta and alphabeta T lymphocytes were determined by flow cytometry of cells stained with monoclonal antibodies to gammadelta and alphabeta T lymphocyte receptors. The activation of these cells was evaluated by flow cytometry of cells stained with monoclonal antibodies to CD69 and HLA-DR. Serial counts and activation of gammadelta and alphabeta T lymphocytes were also determined in eight trauma patients (Injury Severity Score of 31.0 +/- 13.5) during a 2-week observation period. The count of gammadelta T lymphocytes in the peripheral blood of SIRS patients (30.1 +/- 6.0/microL) was significantly lower (P0.05) than that of the healthy volunteers (104.3 +/- 10.9/microL). The expression of CD69, an index of early activation of T lymphocytes, was significantly greater on gammadelta T lymphocytes from SIRS patients (patients 23.9% +/- 3.4%, healthy controls 4.8% +/- 0.6%, P0.05). In trauma patients, the expression of CD69 on gammadelta T lymphocytes increased rapidly within 48 h after injuries. In conclusion, gammadelta T lymphocytes are activated and decreased in the peripheral blood of severe SIRS patients. In trauma patients, the activation of gammadelta T lymphocytes occurs in the fairly acute phase after injuries. These results suggest a significant role for gammadelta T lymphocytes as early responders after severe insult.

Background The gut is an important target organ for injury after severe insult, and resolution of feeding intolerance is crucial for critically ill patients. We investigated gut flora and motility to evaluate the impact of gastrointestinal dysmotility on septic complications in patients with severe systemic inflammatory response syndrome (SIRS). Methods Sixty-three ICU patients with severe SIRS were divided into two groups depending on their intestinal condition. Patients with feeding intolerance comprised patients who had feeding intolerance, defined as ≥300 mL reflux from nasal gastric feeding tube in 24 h, and patients without feeding intolerance comprised patients with no feeding intolerance. We compared fecal microflora, incidences of bacteremia, and mortality between these groups. Key Results Analysis of feces showed that patients with feeding intolerance had significantly lower numbers of total obligate anaerobes including Bacteroidaceae and Bifidobacterium, higher numbers of Staphylococcus, lower concentrations of acetic acid and propionic acid, and higher concentrations of succinic acid and lactic acid than those in patients without feeding intolerance (P ≤ 0.05). Patients with feeding intolerance had higher incidences of bacteremia (86%vs 18%) and mortality (64%vs 20%) than did patients without feeding intolerance (P ≤ 0.05). Conclusions & Inferences Gut flora and organic acids were significantly altered in patients with severe SIRS complicated by gastrointestinal dysmotility, which was associated with higher septic mortality in SIRS patients.

Introduction: Sepsis is a serious medical condition that requires rapidly administered, appropriate antibiotic treatment. Conventional methods take three or more days for final pathogen identification and antimicrobial susceptibility testing. We organized a prospective observational multicenter study in three study sites to evaluate the diagnostic accuracy and potential clinical utility of the SeptiFast system, a multiplex pathogen detection system used in the clinical setting to support early diagnosis of bloodstream infections. Methods: A total of 212 patients, suspected of having systemic inflammatory response syndrome (SIRS) caused by bacterial or fungal infection, were enrolled in the study. From these patients, 407 blood samples were taken and blood culture analysis was performed to identify pathogens. Whole blood was also collected for DNA Detection Kit analysis immediately after its collection for blood culture. The results of the DNA Detection Kit, blood culture and other culture tests were compared. The chosen antimicrobial treatment in patients whose samples tested positive in the DNA Detection Kit and/or blood culture analysis was examined to evaluate the effect of concomitant antibiotic exposure on the results of these analyses. Results: SeptiFast analysis gave a positive result for 55 samples, while 43 samples were positive in blood culture analysis. The DNA Detection Kit identified a pathogen in 11.3% (45/400) of the samples, compared to 8.0% (32/400) by blood culture analysis. Twenty-three pathogens were detected by SeptiFast only; conversely, this system missed five episodes of clinically significant bacteremia (Methicillin-resistant Staphylococcus aureus (MRSA), 2; Pseudomonas aeruginosa ,1 ;Klebsiella spp ,1 ;Enterococcus faecium, 1). The number of samples that tested positive was significantly increased by combining the result of the blood culture analysis with those of the DNA Detection Kit analysis (P = 0.01). Among antibiotic pre-treated patients (prevalence, 72%), SeptiFast analysis detected more bacteria/fungi, and was less influenced by antibiotic exposure, compared with blood culture analysis (P = 0.02). Conclusions: This rapid multiplex pathogen detection system complemented traditional culture-based methods and offered some added diagnostic value for the timely detection of causative pathogens, particularly in antibiotic pre-treated patients. Adequately designed intervention studies are needed to prove its clinical effectiveness in improving appropriate antibiotic selection and patient outcomes.

The gut is an important target organ for stress caused by severe insults such as sepsis, trauma, burn, shock, bleeding and infection. Severe insult to the gut is considered to have an important role in promoting infectious complications and multiple organ dysfunction syndrome. These are sequelae of interactions between deteriorated intestinal epithelium, the immune system and commensal bacteria. The gut is the “motor” of multiple organ failure, and now it is recognized that gut dysfunction is a causative factor in disease progression. The gut flora and environment are significantly altered in critically ill patients, and the number of obligate anaerobes is associated with prognosis. Synbiotic therapy is a combination of probiotics and prebiotics. Probiotic, prebiotic and synbiotic treatment has been shown to be a promising therapy to maintain and repair the gut microbiota and gut environment. In the critically ill, such as major abdominal surgery, trauma and ICU patients, synbiotic therapy has been shown to significantly reduce septic complications. Further basic and clinical research would clarify the underlying mechanisms of the therapeutic effect of probiotic/synbiotic treatment and define the appropriate conditions for use.