Molecular mechanisms of cross-inhibition between nicotinic acetylcholine receptors and P2X receptors in myenteric neurons and HEK-293 cells.

BACKGROUND: P2X(2) and nicotinic acetylcholine receptors (nAChRs) mediate fast synaptic excitation in the enteric nervous system. P2X receptors and nAChRs are functionally linked. This study examined the mechanisms responsible for interactions between P2X2 and alpha3beta4subunit-containing nAChRs. METHODS: The function of P2X2 and alpha3beta4 nAChRs expressed by HEK-293 cells and guinea pig ileum myenteric neurons in culture was studied using whole-cell patch clamp techniques. KEY RESULTS: In HEK-293 cells expressing alpha3beta4 nAChRs and P2X2 receptors, co-application of ATP and acetylcholine caused inward currents that were 56 +/- 7% of the current that should occur if these channels functioned independently (P < 0.05, n = 9); we call this interaction cross-inhibition. Cross-inhibition did not occur in HEK-293 cells expressing alpha3beta4 nAChRs and a C-terminal tail truncated P2X2 receptor (P2X2TR) (P > 0.05, n = 8). Intracellular application of the C-terminal tail of the P2X2 receptor blocked nAChR-P2X receptor cross-inhibition in HEK-293 cells and myenteric neurons. In the absence of ATP, P2X2 receptors constitutively inhibited nAChR currents in HEK-293 cells expressing both receptors. Constitutive inhibition did not occur in HEK-293 cells expressing alpha3beta4 nAChRs transfected with P2X2TR. Currents caused by low (< or =30 micromol L(-1)), but not high (> =100 micromol L(-1)) concentrations of ATP in cells expressing P2X2 receptors were inhibited by co-expression with alpha3beta4 nAChRs. CONCLUSIONS & INFERENCES: The C-terminal tail of P2X2 receptors mediates cross-inhibition between alpha3beta4 nAChR-P2X2 receptors. The closed state of P2X2 receptors and nAChRs can also cause cross-inhibition. These interactions may modulate transmission at enteric synapses that use ATP and acetylcholine as co-transmitters.

[Port stapling for simplified port implantation in laparoscopic adjustable gastric banding]. / Port-Stapling zur Erleichterung der Portimplantation bei der laparoskopischen Magenbandoperation.

BACKGROUND: The purpose of this study was to evaluate the applicability of a port stapling device to simplify and improve port implantation in laparoscopic adjustable gastric banding (LAGB). METHODS: From November 2005 to September 2006, a prospective study was conducted on 23 consecutive patients who underwent LAGB with Swedish adjustable gastric banding. Patients were randomized to either conventional titanium-port implantation or port stapling using the "Velocity" device. RESULTS: No differences in age, body weight, body mass index, fascia depth or incision length were reported between the groups. Port implantation time was significantly less using port stapling (90+/-24 s) compared to conventional port implantation (521+/-138 s). Port related complaints postoperatively and at follow-up were equal in both groups. CONCLUSIONS: Port stapling is an excellent tool to facilitate port implantation, particularly in massively obese patients with a thick abdominal wall.

Reduced liver apoptosis after venous systemic oxygen persufflation in non-heart-beating donors.

Graft injury caused by warm ischemia in livers from non-heart-beating donors (NHBDs) strongly affects posttransplantation outcome and is associated with liver apoptosis, which is mediated by death receptors, such as Fas, a surface receptor of the tumor necrosis factor (TNF)-alpha family. The aim of this study was to test the ability of venous systemic oxygen persufflation (VSOP) to reduce apoptotic changes and Fas activation in the liver after warm ischemic insult in vivo. Livers of male Wistar rats were harvested 30 min after cardiac arrest from non-heart-beating donors (NHBD) with (NHBD + O2) or without (NHBD) application of gaseous oxygen during the cold storage period via the suprahepatic caval vein. After 24 h of storage in University of Wisconsin solution at 4 degrees C, viability of the livers was assessed upon isolated reperfusion in vitro. Conventional signs of tissue damage like enzyme release and bile production showed a significantly elevated nonspecific cell injury in the NHBD group. TUNEL staining revealed increased DNA fragmentation of sinusoidal endothelial cells in the NHBD group and more apoptotic hepatocytes than in the control group. All these alterations could be almost abrogated by the use of VSOP in the NHBD + O2 group. The immunohistochemical staining of Fas antigen expression showed a significantly elevated Fas receptor expression in the NHBD and NHBD + O2 groups, in accord with an eightfold increase of Fas receptor mRNA detected by real-time reverse-transcription polymerase chain reaction (RT-PCR). These results demonstrate that the postischemic apoptotic rate of sinusoidal endothelial cells in NHBD livers can be reduced by the use of VSOP. A significant improvement in liver integrity and viability was obtained with this technique, without influencing the expression of Fas expression.

[Massive localized lymphedema - a rare entity]. / Massives lokalisiertes Lymphödem - eine seltene Entität.

Massive localized lymphedema is a rare disease. Only a few cases have been described in the literature. These monstrous pseudotumors of the subcutis are mostly localized in the inguinal region or at the lower extremity. These tumors often show a slow growth for many years. Besides hernias, lipomatous tumors must be distinguished. The therapy of choice is the excision of the tumor. Relapse is not uncommon in the few cases described in the literature so far. The diagnostic procedure and therapy of an 48-year-old women with a massive localized lymphedema weighing about 22 kg are demonstrated and discussed.

Changes in TH1/TH2 immunity after endovascular and conventional infrarenal aortic aneurysm repair: its relevance for clinical practice.

OBJECTIVE: to evaluate local surgical trauma induced by endovascular (TPEG) and conventional infrarenal aortic aneurysm repair (AAA-C), the inflammatory response and changes in cell-mediated and antibody-mediated immunity as illustrated by the type-1/type-2 T-helper (TH1/TH2) cell balance were investigated. DESIGN: prospective study. PATIENTS AND METHODS: sixteen patients were included, eight patients underwent AAA-C and eight TPEG. Venous peripheral blood samples were collected 24h preoperatively and 24, 48, 72h, 5 and 7 days postoperatively. Besides the WBC, intracellular TH1/TH2 cytokines (IFN-gamma/IL-4) and the cell surface markers HLA-DR on monocytes and CD23 on B cells were measured by four colour flow cytometry. RESULTS: statistically significant higher values in the AAA-C group were demonstrated for neutrophiles. The TH1/TH2 immunobalance (expressed by forming the ratio of IFN-(gamma/IL-4 producing T cells as well as by the ratio of HLA-DR(pos) monocytes/CD23(pos) B-cells) showed a significant shift towards TH2 immunity in the AAA-C group whereas TPEG led to a significant lesser shift 24-72h after surgery (p < 0.05). CONCLUSIONS: TPEG leads to a minor distortion of the TH1/TH2 immunobalance. This implies that TPEG is a less stressing procedure, that is especially beneficial in patients whose conditions are considered less suitable for AAA-C due to age and serious comorbidity.

Practical fast gas chromatography for contract laboratory program pesticide analyses.

An approach to shortening the analysis time for practical fast gas chromatography (GC) by using Method Translator software, which can be downloaded free from the Internet, is presented. This software simplifies the process of optimizing temperature programming while changing column dimensions, carrier gas type, and flow. Basic chromatographic theory is employed in a practical manner for adjusting column dimensions for optimal performance. In addition, electronic pneumatic control and high oven ramp rates make it easier to achieve fast analysis times without reproducibility problems. This practical approach is demonstrated using Contract Laboratory Program pesticide analytes. The factors found to be most important in decreasing the analysis time without a loss of performance are utilization of GC columns having smaller diameters and substitution of hydrogen for helium as the carrier gas.

Increased susceptibility to apoptosis in circulating lymphocytes of critically ill patients.

BACKGROUND AND AIMS: Lymphocyte apoptosis may influence immune responsiveness in systemic inflammation. Therefore, we investigated whether early signs of apoptosis (i.e., annexin-V binding and cell shrinkage) in peripheral lymphocytes were different among patients with severe sepsis, critically ill, nonseptic patients after major surgery, and healthy individuals. PATIENTS/METHODS: Ten patients with severe sepsis and ten critically ill, nonseptic patients after major surgery admitted to a surgical intensive care unit in a university hospital were included in the study. In addition, ten healthy blood donors were included for comparison. We investigated early signs of apoptosis using flow cytometric measurement of annexin-V binding to the cell surface and cell shrinkage of peripheral lymphocytes. RESULTS: The percentage of apoptotic lymphocytes determined as annexin-V positive and propidium iodide negative cells was increased in freshly prepared cells of patients with severe sepsis (11.4 +/- 0.5%) and critically ill, nonseptic patients after major surgery (18.5 +/- 2.0%) relative to healthy blood donors (4.4 +/- 0.5%) (P < 0.05). No significant difference between patients with severe sepsis and patients after major surgery were found. Annexin-V binding increased significantly after OKT-3 stimulation of lymphocytes in patients with severe sepsis (34.4 +/- 1.6%), patients after major surgery (33.8 +/- 3.4%), and healthy blood donors (21.1 +/- 2.8%). No significant difference among groups was detected following OKT-3 stimulation. Furthermore, freshly isolated peripheral lymphocytes of patients with severe sepsis and critically ill, nonseptic patients after major surgery revealed a significantly higher proportion of cell shrinkage than in healthy blood donors (55.0 +/- 2.2%, 21.5 +/- 2.4% vs 3.6 +/- 0.7%; P < 0.05). CONCLUSION: Circulating lymphocytes of critically ill patients show a high degree of early signs of cellular apoptosis. This may contribute to hyporesponsiveness of immune cells in systemic inflammation.

Use of the Polyflex stent in the palliative therapy of esophageal carcinoma: results in 14 cases and review of the literature.

BACKGROUND: Several prospective randomized trials have shown that self-expanding stents have advantages over conventional plastic tubes. Nevertheless, the optimal stent has not yet been developed. The Polyflex stent is a completely new model that represents an improvement over the old metal stents. We have used this stent in a prospective study and herein our present preliminary results. METHODS: In 14 patients with nonresectable esophageal carcinoma, the Polyflex stent was implanted to reduce dysphagia. The grade of dysphagia, the complications following intervention, and the patients' total survival time were documented prospectively every 4 weeks. RESULTS: The implantation of the stent was successful in all cases. The grade of the dysphagia was reduced from 3.0 to 0.5 after stent implantation. One patient died during the hospital stay from a non-stent-induced complication. Stent dislocation occurred once, and tumor overgrowth at the stent margins was observed twice. The mean survival time was 6.2 months, and the reintervention rate was 21.3%. CONCLUSION: The new Polyflex stent, which is based on a completely new design, can be implanted without any difficulty and has had very good short- and long-term results. Therefore, it is a worthy alternative to the metal stents in current use.

Detection of circulating epithelial cells after surgery for benign breast disease.

BACKGROUND: Cytokeratins are predominantly expressed in epithelial cells and their malignant counterparts. Ultrasensitive methods for cytokeratin messenger RNAs (mRNAs) can detect rare circulating tumor cells consistent with hematogenous dissemination in epithelial-derived malignancies, including breast carcinomas. Intraoperative tumor-cell shedding may contribute to this process; this hypothesis is based on the assumption that only tumor cells can be mobilized during surgical manipulation. METHODS AND RESULTS: The present study addresses this issue by using cytokeratin 19 mRNA detection by reverse transcription-polymerase chain reaction (RT-PCR) in preoperative and postoperative blood samples from 54 patients undergoing excisional biopsy for benign breast disease; 22 healthy volunteers represented the control group. No cytokeratin RT-PCR positivity was found in the control or preoperative samples. Cytokeratin RT-PCR positivity was found in 21 postoperative samples (39%). CONCLUSIONS: This finding shows that benign epithelial cells can be mobilized during breast surgery; this effect of surgical manipulation warrants caution in the interpretation of RT-PCR positivity for cytokeratin mRNA in the peripheral blood of patients undergoing surgery for breast cancer.

Myelodysplastic syndrome at a large tertiary care community hospital: analysis according to the international prognostic scoring system.

The outcome of patients diagnosed myelodysplastic syndromes (MDS) between 1990 and 1997 from William Beaumont Hospital (WBH) was analyzed according to the International Prognostic Scoring System (IPSS) risk categorization. A retrospective study of 195 MDS patients wa s performed. Seventy-nine patients with MDS, in whom a karyotype was obtained and with an adequate follow-up were included in the final analysis. Cases of proliferative CMML (WBC > 12x10(9)/l) were excluded from the study. The overall median survival was 3.1 years, and median survival stratified by IPSS was 3.4, 4.1 and 0.5 years for the INT-1, INT-2 and high risk group and not yet reached for the low risk group. The overall survival by IPSS subcategorization were 6.88, 5.29, 5.30 and 2.12 years for the low, INT-1, INT-2, and high risk groups respectively. Cytogenetics were significant in predicting the overall survival. The IPSS score stratified patients into risk categories for development of AML. The risk of development into AML was 8, 8, 33 and 54% for the low, INT-1, INT-2 and high risk groups, respectively. We conclude that IPSS score can be useful in predicting survival and AML evolution in some MDS patients.

RIBP, a novel Rlk/Txk- and itk-binding adaptor protein that regulates T cell activation.

A novel T cell-specific adaptor protein, RIBP, was identified based on its ability to bind Rlk/Txk in a yeast two-hybrid screen of a mouse T cell lymphoma library. RIBP was also found to interact with a related member of the Tec family of tyrosine kinases, Itk. Expression of RIBP is restricted to T and natural killer cells and is upregulated substantially after T cell activation. RIBP-disrupted knockout mice displayed apparently normal T cell development. However, proliferation of RIBP-deficient T cells in response to T cell receptor (TCR)-mediated activation was significantly impaired. Furthermore, these activated T cells were defective in the production of interleukin (IL)-2 and interferon gamma, but not IL-4. These data suggest that RIBP plays an important role in TCR-mediated signal transduction pathways and that its binding to Itk and Rlk/Txk may regulate T cell differentiation.

Impaired inducibility of heat shock protein 70 in peripheral blood lymphocytes of patients with severe sepsis.

OBJECTIVE: To determine the extent of the potentially protective heat shock protein 70 response in peripheral blood lymphocytes of patients with severe sepsis after ex vivo lipopolysaccharide stimulation. DESIGN: Entry study of consecutive patients with severe sepsis, those who were critically ill or nonseptic after major surgery, and healthy blood donors. SETTING: Surgical intensive care unit in a university hospital. PATIENTS: Ten patients with diagnoses of severe sepsis; ten critically ill, nonseptic patients after major surgery; and ten healthy blood donors. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: We investigated the ex vivo endotoxin-inducible expression of heat shock protein 70 in peripheral blood lymphocytes of patients with severe sepsis by means of flow cytometry. Only negligible amounts of inducible intracellular heat shock protein 70 accumulation (<4.2% of lymphocytes) could be detected in peripheral blood lymphocytes without lipopolysaccharide stimulation. The proportion of cells accumulating heat shock protein 70 after treatment with lipopolysaccharide was distinctly lower in patients with severe sepsis (p < .05) than in critically ill, nonseptic patients after major surgery and healthy blood donors (38.3+/-3.3%, 82.2+/-4.5%, and 70.9+/-3.9%, respectively; mean +/- SEM; n = 10). Patients with clinical signs of recovery from severe sepsis showed an increase in heat shock protein 70 expression. CONCLUSIONS: Inducibility of ex vivo heat shock protein 70 was impaired in peripheral blood lymphocytes of patients with severe sepsis. The impaired expression of the potentially protective heat shock protein 70 may contribute in vivo to immune dysfunction, because intact functioning of T and B lymphocyte responses is of central importance in resisting infection in severe sepsis. Monitoring of inducible heat shock protein 70 in peripheral blood lymphocytes may contribute to the evaluation of the immune consequences of severe sepsis.

Impact of clinical, pathologic, and treatment-related factors on outcome in patients with locally advanced breast cancer treated with multimodality therapy.

The authors reviewed the experience at their institution treating patients with locally advanced breast cancer using multimodality therapy to identify clinical, pathologic, and treatment-related factors affecting outcome. One hundred patients with locally advanced breast cancer were treated with definitive therapy at William Beaumont Hospital. Three patients had stage IIB disease, 45 patients had stage IIIA disease, and 52 patients had IIIB disease. Thirteen patients had inflammatory breast carcinoma. Seventy-four patients (74%) received trimodality therapy consisting of systemic therapy, radiation therapy, and surgery. Systemic therapy was delivered to 90 patients. Eighty-three patients (83%) received adjuvant radiation therapy. Eighty-five patients underwent mastectomy (85%). Multiple clinical, pathologic, and treatment-related factors were analyzed for their impact on outcome. The median follow-up was 47 months. Overall, the 5-year actuarial rates of local control, disease-free survival, overall survival, and cause-specific survival were 81%, 43%, 53%, and 55%, respectively. The 5-year actuarial cause-specific survival rates for patients with inflammatory breast carcinoma, stage IIIA disease, and stage IIIB disease were 25%, 55%, and 53%, respectively. On multivariate analysis, local control was improved with radiation therapy (p = 0.008) and the absence of inflammatory breast carcinoma (p = 0.008). Disease-free survival was improved with the addition of radiation therapy (p = 0.001) and with less than four positive lymph nodes (p = 0.003). Distant metastasis-free survival was improved in patients without inflammatory breast carcinoma (p = 0.0249) and with less than four involved lymph nodes (p = 0.0135). Cause-specific survival and overall survival were adversely affected by the presence of inflammatory breast carcinoma (p = 0.0135 and p = 0.0325, respectively) or four or more involved lymph nodes (p = 0.0082 and p = 0.012, respectively). Radiation therapy appears to be a critical component in the overall treatment of patients with locally advanced breast cancer by improving the rates of local control and disease-free survival. Other adverse factors for survival include four or more positive lymph nodes and inflammatory breast carcinoma.

Benign and malignant cells in effusions: diagnostic value of image DNA cytometry in comparison to cytological analysis.

Identifying tumor cells in body cavity fluids reliably is a well-known diagnostic problem. Since cytometric quantitation of nuclear DNA content appears to be a promising new tool in the diagnosis and prognostic evaluation of many solid human tumors, we examined its validity in detecting malignant cells in cytologically positive effusions. For this purpose, image DNA cytometric measurements, including the evaluation of DNA-ploidy and the calculation of the DNA index (DI), were performed in 80 body cavity fluids. The results were correlated with cytology, clinical course and final histological diagnoses. We used aneuploidy, as shown by interactive image DNA cytometry, as a marker for the malignancy of cells that occur in body cavity fluids with a 100% specificity and 94.8% sensitivity. Cytological investigation showed a 92.3% specificity and 95.4% sensitivity. Combining both methods raised the specificity to 100% and the sensitivity to 98.5% and had a positive predictive value of 100% and a negative predictive value of 93.8%. The DNA-index (DI) was significantly higher in malignant effusions than in benign effusions: 1.5 +/- 0.74 (mean +/- SD) versus 1.11 +/- 0.26 (p < 0.05). Along with the difficult cytological evaluation of malignant cells in body cavity fluids, image DNA cytometry can be a helpful additional method for evaluating these cells. Combining the two techniques results in a highly specific and sensitive prediction of malignant cells. We, therefore, suggest using these methods for the reliable identification of tumor cells in effusions.

[Increased CA-19-9 caused by a splenic cyst: a rare differential diagnosis]. / CA19-9-Erhöhung durch eine Milzzyste: Eine seltene Differentialdiagnose.

Adenocarcinoma of the gastrointestinal tract, particularly primary adenocarcinoma of the pancreas lead to elevation of serum carbohydrate 19-9 (CA19-9) levels. Some benign disorders, like inflammatory diseases also cause CA19-9 tumor marker elevation. An extremely rare entity are benign splenic cysts which express this tumormarker as reported here.

[Palliative therapy of esophageal carcinoma]. / Palliative Therapie des Osophaguskarzinoms.

For palliative endoscopic treatment of malignant dysphagia many different techniques are practiced. The differentiated usage of all techniques depending on the characteristics of the tumor leads to a good result with favorable costs. The following differentiated palliative regimen is presented: in the cervical part of the esophagus laser therapy or stent implantation are used, depending on the type of tumor growth. In the center section and the distal part of the esophagus usually palliative endoscopic pertubation is performed. If the stenosis is very soft and short a stent is implanted as alternative. In the gastroesophageal part flexible stents achieve the best results. Alternative laser therapy can be used.

[The employment schedule act from the viewpoint of the ordinary surgeon]. / Das Arbeitszeitgesetz aus der Sicht des nicht-leitenden Chirurgen.

In Germany, the law of labor time was instituted in hospitals on January 1st, 1996 to regulate working hours, times of rest, breaks, and Sunday and holiday work. Problems in the realization of this law arise in daily practice, especially with regard to maximum working hours and post on-call times of rest. The compliance with the law would necessitate an enormous increase in new jobs for physicians, which is associated with an unfinanceable rise in costs. Solutions are either a reduction in performance or the omission to document the productivity. Therefore, surgeons in salaried employment in German hospitals constantly face the conflict of legality and legitimacy.

[Changes in the cytokine concentration (Il-6, Il-8, Il-1ra) and their cellular expression of membrane molecules (CD25, CD30, HLA-DR) after surgical trauma]. / Veränderung der Zytokinkonzentration (IL-6, IL-8, IL-1 RA) und der zellulären Expression von Membranmolekülen (CD25, CD30, HLA-DR) nach operativem Trauma.

UNLABELLED: Elective surgical approaches and trauma cause changes in the production of different cytokines, an increased production of acute phase protein and changes in the expression of different cell surface markers. METHODS: In a prospective study we examined the C-reactive protein level, the production of the cytokines IL-6, IL-8 and IL-1 RA in 25 laparoscopic and 21 conventional cholecystectomies. In addition the cell surface markers CD25 and CD30 on different cell populations and HLA-DR on monocytes were measured. Statistical analysis was made by Student's-t-test and Mann-Whitney's rank sum test. RESULTS: The humoral markers showed a more distinct increase in patients operated on conventionally two and 24 hours after surgery, the differences between the two surgical approaches were significant (p < 0.01). The cell surface markers CD25 and CD30 showed the same reaction. The HLA-DR expression on monocytes was significantly lower in patients operated on conventionally. CONCLUSIONS: Elective surgical approaches cause changes in the immune system, which can be evaluated by the reaction of cytokines and cell surface markers. Laparoscopic cholecystectomies cause less activation of the immune system than conventional operations.

[Women in surgery--equal surgeons]. / Frauen in der Chirurgie--Ebenbürtige Chirurgen?

In the specialty "surgery" women are underrepresented, especially in leading positions. The reason for that is not less motivation, less ability or less capacity of women, but the working conditions. For women in surgery working conditions are so unfavourable that only few women make their career in surgery.

Analytical and numerical techniques for the evaluation of free radical damage in cultured cells using scanning laser microscopy.

Scanning laser microscopy (SLM) was used to develop an assay to visualize the generation of intracellular reactive oxygen species (ROS) and to evaluate the effect of the lipophilic antioxidant U-87,663 on ROS formation. Cultured N18 neuroglioma cells were challenged by extracellular addition of cumene hydroperoxide, and subsequent intracellular generation of ROS was characterized by measuring the fluorescence intensity of the ROS indicator 2',7'-dichlorofluorescein (DCF). The kinetics of the reaction between ROS and the indicator DCF, or the antioxidant U-87,663, can be most accurately assessed if results from individual cell clusters are analyzed independently. It is possible and necessary to account for the these experimental and analytical properties in order to characterize the properties of the antioxidant activity precisely. We determined that the temporal increase in DCF fluorescence was consistent with the reaction of DCF with free radicals generated from cumene hydroperoxide, as was the loss of fluorescence from U-87,663. The rate constants for the free radical reactions revealed that ROS reaction with DCF is 10 times faster than with U-87,663. These differences in reaction rates combined with differences in the cellular distribution of the ROS indicator DCF, the antioxidant U-87,663, and the bulk of the ROS prevented detection of any protection of U-87,663 may offer.