Fresh tissue procurement and preparation for multicompartment and multimodal analysis of the prostate tumor microenvironment.

BACKGROUND: Prostatic cancers include a diverse microenvironment of tumor cells, cancer-associated fibroblasts, and immune components. This tumor microenvironment (TME) is a known driving force of tumor survival after treatment, but the standard-of-care tissue freezing or fixation in pathology practice limit the use of available approaches/tools to study the TME's functionality in tumor resistance. Thus, there is a need for approaches that satisfy both clinical and laboratory endpoints for TME study. Here we present methods for clinical case identification, tissue processing, and analytical workflow that are compatible with standard histopathology while enabling molecular and functional interrogation of prostate TME components. METHODS: We first performed a small retrospective review to identify cases where submission of alternate prostate tissue slices and a parallel live tissue processing protocol complement traditional histopathology and enable viable multicompartment analysis of the TME. Then, we tested its compatibility with commonly employed methods to study the microenvironment including quantification of components both in situ and after tissue dissociation. We also evaluated tissue digestion conditions and cell isolation techniques to aid various molecular and functional endpoints. RESULTS: We identified Gleason Grade Group 3+ clinical cases where tumor volume was sufficient to allow slicing of unfixed tissue and distribution of alternating tissue slices to standard-of-care histopathology and viable multi-modal TME analyses. No single method was found that preserved cellular sub-types for all downstream readouts; instead, tissues were further divided so techniques could be catered to each endpoint. For instance, we show that incorporating the protease dispase into tissue dissociation improves viability for culture and functional analyses but hinders immune cell analysis by flow cytometry. We also found that flow activated cell sorting provides highly pure cell populations for quantitative reverse-transcription polymerase chain reaction and RNA-seq while isolation using antibody-labeled paramagnetic particles facilitated functional coculture experiments. CONCLUSIONS: The identification of candidate cases and use of these techniques enable translational research and the development of molecular and functional assays to facilitate prostate TME study without compromising standard-of-care histopathological diagnosis. This allows bridging clinical histopathology and further interrogation of the prostate TME and promises to advance our understanding of tumor biology and unveil new predictive and prognostic markers of prostate cancer progression.

Engineering Epithelial-Mesenchymal Microtissues to Study Cell-Cell Interactions in Development.

Intercellular signaling drives human development, but there is a paucity of in vitro models that recapitulate important tissue architecture while remaining operationally simple and scalable. As an example, formation of the upper lip and palate requires the orchestrated proliferation and fusion of embryonic facial growth centers and is dependent on paracrine epithelial-mesenchymal signaling through multiple pathways including the Sonic Hedgehog (SHH), transforming growth factor-beta (Tgf-ß), bone morphogenic protein (BMP), and epidermal growth factor (EGF) pathways. We have developed a robust, throughput-compatible microphysiological system to model intercellular signaling including epithelial-mesenchymal interactions that is useful for studying both normal and abnormal orofacial development. We describe the construction and operation of an engineered microplate created using CNC micromilling of 96-well microtiter plates capable of containing up to 20 epithelial-mesenchymal microtissues. A dense three-dimensional mesenchyme is created by embedding cells (O9-1, 3T3) in a biomimetic hydrogel. An epithelial layer is then overlayed on the microtissue by loading cells in engineered microchannels that flank the microtissue. The result is an engineering epithelial-mesenchymal interface that is both on and perpendicular to the imaging plane making it suitable for high-content imaging and analysis. The resulting microtissues and device are compatible with diverse analytical techniques including fluorescent and luminescent cell health and enzymatic reporter assays, gene expression analyses, and protein staining. This tractable model and approach promise to shed light on critical processes in intercellular signaling events in orofacial development and beyond.

Microfluidic tumor-on-a-chip model to evaluate the role of tumor environmental stress on NK cell exhaustion.

Solid tumors generate a suppressive environment that imposes an overwhelming burden on the immune system. Nutrient depletion, waste product accumulation, hypoxia, and pH acidification severely compromise the capacity of effector immune cells such as T and natural killer (NK) cells to destroy cancer cells. However, the specific molecular mechanisms driving immune suppression, as well as the capacity of immune cells to adapt to the suppressive environment, are not completely understood. Thus, here, we used an in vitro microfluidic tumor-on-a-chip platform to evaluate how NK cells respond to the tumor-induced suppressive environment. The results demonstrated that the suppressive environment created by the tumor gradually eroded NK cell cytotoxic capacity, leading to compromised NK cell surveillance and tumor tolerance. Further, NK cell exhaustion persisted for an extended period of time after removing NK cells from the microfluidic platform. Last, the addition of checkpoint inhibitors and immunomodulatory agents alleviated NK cell exhaustion.

A Microphysiological Approach to Evaluate Effectors of Intercellular Hedgehog Signaling in Development.

Paracrine signaling in the tissue microenvironment is a central mediator of morphogenesis, and modeling this dynamic intercellular activity in vitro is critical to understanding normal and abnormal development. For example, Sonic Hedgehog (Shh) signaling is a conserved mechanism involved in multiple developmental processes and strongly linked to human birth defects including orofacial clefts of the lip and palate. SHH ligand produced, processed, and secreted from the epithelial ectoderm is shuttled through the extracellular matrix where it binds mesenchymal receptors, establishing a gradient of transcriptional response that drives orofacial morphogenesis. In humans, complex interactions of genetic predispositions and environmental insults acting on diverse molecular targets are thought to underlie orofacial cleft etiology. Consequently, there is a need for tractable in vitro approaches that model this complex cellular and environmental interplay and are sensitive to disruption across the multistep signaling cascade. We developed a microplate-based device that supports an epithelium directly overlaid onto an extracellular matrix-embedded mesenchyme, mimicking the basic tissue architecture of developing orofacial tissues. SHH ligand produced from the epithelium generated a gradient of SHH-driven transcription in the adjacent mesenchyme, recapitulating the gradient of pathway activity observed in vivo. Shh pathway activation was antagonized by small molecule inhibitors of epithelial secretory, extracellular matrix transport, and mesenchymal sensing targets, supporting the use of this approach in high-content chemical screening of the complete Shh pathway. Together, these findings demonstrate a novel and practical microphysiological model with broad utility for investigating epithelial-mesenchymal interactions and environmental signaling disruptions in development.

Vital ex vivo tissue labeling and pathology-guided micropunching to characterize cellular heterogeneity in the tissue microenvironment.

Cellular heterogeneity within the tissue microenvironment may underlie chemotherapeutic resistance and response, enabling tumor evolution; however, this heterogeneity it is difficult to characterize. Here, we present a new approach-pathology-guided micropunching (PGM)-that enables identification and characterization of heterogeneous foci identified in viable human and animal model tissue slices. This technique consists of live-cell tissue labeling using fluorescent antibodies/small molecules to identify heterogeneous foci (e.g., immune infiltrates or cells with high levels of reactive oxygen species) in viable tissues, coupled with a micropunch step to isolate cells from these heterogeneous foci for downstream molecular or vital functional analysis. Micropunches obtained from epithelial or stromal fibroblast foci in human prostate tissue show 6- to 12-fold enrichment in transcripts specific for EpCam/cytokeratin 8 and vimentin/a-smooth muscle actin/integrin 1-α, respectively. Transcriptional enrichment efficiency agrees with epithelial and stromal laser capture microdissection samples isolated from human prostate. Micropunched foci show a loss of cellular viability in the periphery, but centrally localized cells retained viability before and after dissociation and grew out in culture.

The Aryl Hydrocarbon Receptor is a Repressor of Inflammation-associated Colorectal Tumorigenesis in Mouse.

OBJECTIVE: To determine the role of the aryl hydrocarbon receptor (AHR) in colitis-associated colorectal tumorigenesis. BACKGROUND: Colorectal cancer (CRC) is the third most commonly diagnosed cancer in United States. Chronic intestinal inflammation increases the risk for the development of CRC. We investigated the involvement of AHR, a ligand-activated transcriptional regulator, in colitis-associated colorectal tumorigenesis. METHODS: We used a mouse model of colitis-associated colorectal tumorigenesis that employs treatment with azoxymethane and dextran sodium sulfate. We examined the role of AHR using both an Ahr-deletion mouse model (Ahr) and treatment with the AHR pro-agonist indole-3-carbinol (I3C). Incidence, multiplicity, and location of tumors were visually counted. Tumors were defined as neoplasms. Intestinal inflammation was assessed by quantitative PCR for proinflammatory markers and colon length. Data were evaluated and compared using GraphPad Prism software (version 6, La Jolla, CA). RESULTS: Tumor incidence was increased 32% in Ahr null mice and tumor multiplicity was approximately increased 3-fold compared with wild-type mice (2.4 vs 7; P < 0.05). Furthermore, tumor multiplicity was reduced 92% by treatment of I3C in wild-type mice, whereas the suppressor effect of I3C was not observed in Ahr null mice (P < 0.05). CONCLUSIONS: We found that AHR plays a protective role in colitis-associated colorectal tumorigenesis. This conclusion is based on the observations that Ahr null mice showed increased number of colorectal tumors, and mice treated with I3C exhibited fewer tumors. This study supports the use of AHR agonists such as I3C as a chemopreventive therapy for IBD-associated CRC in human patients.

Aryl hydrocarbon receptor-dependent apoptotic cell death induced by the flavonoid chrysin in human colorectal cancer cells.

The polyphenolic flavone chrysin has been evaluated as a natural chemopreventive agent due to its anti-cancer effects in a variety of cancer cell lines. However, the mechanism of the chemopreventive effect has been not well established, especially in human colorectal cancer cells. We evaluated the chemopreventive effect of chrysin in three different human colorectal cancer cell lines. We found that chrysin treatment consequently reduced cell viability via induction of apoptosis. We identified that the involvement of up-regulation of pro-apoptotic cytokines tumor necrosis factor (Tnf) α and ß genes and consequent activation of the TNF-mediated transcriptional pathway in chrysin-induced apoptosis. Using our generated AHR siRNA expressing colorectal cancer cells, we demonstrated that the chrysin-induced up-regulation of Tnfα and ß gene expression was dependent on the aryl hydrocarbon receptor (AHR), which is a ligand-receptor for chrysin. Subsequently, we found that the AHR siRNA expressing colorectal cancer cells were resistant to chrysin-induced apoptosis. Therefore, we concluded that AHR is required for the chrysin-induced apoptosis and the up-regulation of Tnfα and ß gene expression in human colorectal cancer cells.

Diabetes: a risk factor for catheter-associated infections.

BACKGROUND AND OBJECTIVES: The incidence of infectious complications associated with continuous regional anesthesia techniques is a matter of concern. Our objective was to determine whether patients suffering from diabetes are at an increased risk of catheter-related infectious complications. METHODS: The German Network for Regional Anaesthesia database was analyzed between 2007 and 2012. After proof of plausibility, data of 36,881 patients undergoing continuous regional anesthesia were grouped in I: no diabetes (n = 32,891) and II: any diabetes (n = 3990). The analysis focused on catheter-related infections after strict definition. Differences among the groups were tested with t and χ tests. Odds ratios were calculated with logistic regression and adjusted for potential confounders. RESULTS: Patients with a diagnosis of diabetes had an increased incidence of catheter-related infections (no diabetes 3.0% vs any diabetes 4.2%; P < 0.001). Among all patients, diabetes remained an independent risk factor for infections for all sites after the adjustment for potential confounders (odds ratio [OR] = 1.26; 95% confidence interval [95% CI], 1.02-1.55; P = 0.036). The risk of infection was significantly increased in peripheral catheters only in the lower limb (adjusted OR = 2.42; 95% CI, 1.05-5.57; P = 0.039). If neuraxial catheters were used, the risk was significantly increased only in lumbar epidural (adjusted OR = 2.09; 95% CI, 1.18-3.73; P = 0.012) for diabetic patients compared with nondiabetic patients. CONCLUSIONS: The presence of diabetes is associated with an increased risk for catheter-related infections in lower limb and lumbar epidural. Specific care should be taken to avoid and detect infections in this population.

Utility of micro-ribonucleic acid profile for predicting recurrence of rectal cancer.

BACKGROUND: In early-stage rectal cancer, the surgeon must decide between the high morbidity of radical surgery and the high recurrence rates of local excision. A prognostic marker could improve patient selection and lower recurrence rates. Micro-ribonucleic acids (miRNAs), small RNAs that often inhibit tumor suppressors, have shown prognostic potential in colorectal cancer. We hypothesized that high miRNA levels in malignant tissue from early-stage rectal cancer patients could predict recurrence after local excision. MATERIALS AND METHODS: We identified 17 early-stage rectal cancer patients treated with local excision between 1990 and 2005, four of whom had recurrences. Total RNA was extracted from benign and malignant tissue and used in quantitative real-time reverse transcriptase polymerase chain reaction to probe for miR-20a, miR-21, miR-106a, miR-181b, and miR-203. MiRNA data were evaluated for association with recurrence using univariate analysis with Wilcoxon rank sum test. RESULTS: Malignant tissue in both patients who had recurrences and patients who did not have recurrences had equivalently high levels of miRNA. However, the benign tissue of patients who recurred contained significantly higher levels of all five miRNAs when compared with the benign tissue of nonrecurrent patients despite having no histological differences. CONCLUSIONS: This is the first study to show that high miRNA levels of histologically benign tissue obtained from the surgical margin of locally excised rectal cancers can predict recurrence. The malignant miRNA levels did not have predictive value. Further investigation of miRNAs is needed to explore their potential for a more accurate prognosis of rectal cancer.

Electrostimulation with or without ultrasound-guidance in interscalene brachial plexus block for shoulder surgery.

In a prospective controlled trial to compare conventional interscalene brachial plexus block (ISBPB) using anatomic landmarks and electro-stimulation with a combined technique of ultrasound guidance followed by nerve stimulation, 60 patients were randomized into 2 matched equal groups: Group A using nerve stimulation (NS) alone and Group B using the combination of ultrasound and NS. The time to detect the plexus (3.9 ± 4 min in Group A and 3.3 ± 1.4 min in Group B) was not significantly different. We needed to reposition the needle once (n = 13) or twice (n = 4) in Group B. First-shot motor response was achieved in all but one patient in Group A; here we were only able to locate the plexus by use of ultrasound. None of the patients needed general anaesthesia. There were no significant differences between postoperative pain, motor power, or patient's satisfaction. ISBPB seems similarly effective using electro-stimulation and ultrasound if performed by experienced anesthesiologists.

Long-term ventilation of patients with Duchenne muscular dystrophy: experiences at the Neuromuscular Centre Ulm.

The various measures used to treat the symptoms of Duchenne muscular dystrophy (DMD), i.e. medication with steroids, early operation on contractures and spine deformities as well as cardiac diagnostics and therapy, should always be accompanied by careful monitoring of the patient's respiratory status. Therapy for respiratory failure, in particular long-term ventilation, is now generally accepted as essential for DMD patients. The provision of assisted ventilation has made a decisive contribution to the quality of life for older patients and the stigma hitherto attached to it as being merely a means of keeping a patient comfortable towards the end of life has now been dispelled. Even outside the hospital, assisted ventilation has become routine. These days it is not uncommon for patients on assisted ventilation to have their life extended by 10 years or more. Non-invasive ventilation is sufficient if used concomitantly with coughing aids. Before undergoing orthopaedic surgery the patient' s respiratory status has to be carefully assessed in order to minimize the risk of perioperative complications. Feeding and swallowing problems may develop if the patient has a scoliosis of the cervical spine region, even if he has had thoraco-lumbar spine surgery. There is still insufficient awareness of this potential problem in relation to respiratory care. Interdisciplinary collaboration between hospitals, general practitioners, muscle and respiratory centres, as well as advocacies and self-help groups is vital. The administration of aids to support DMD patients is now facilitated by guidelines drawn up by several centres of excellence. Here we mainly describe the historic development of respiratory care at the Ulm Neuromuscular Centre.

ZM336372 induces apoptosis associated with phosphorylation of GSK-3beta in pancreatic adenocarcinoma cell lines.

INTRODUCTION: ZM336372 is small molecule tyrosine kinase modulator. It has been shown to inhibit glycogen synthase kinase-3beta (GSK-3beta) through phosphorylation of GSK-3beta at Ser 9. GSK-3beta has previously been shown to mediate cell survival in pancreatic cancer cells. Here we determine the effects of ZM336372 on GSK-3beta phosphorylation, apoptosis, and growth in pancreatic adenocarcinoma cell lines. METHODS: Panc-1 and MiaPaCa-2 cells were treated with ZM336372 or lithium chloride (LiCl) and compared with solvent control. The effects on proliferation for each cell line were determined using the MTT assay. Western blot analysis was performed to examine the effects of treatment on the phosphorylation of GSK-3beta. In addition, western blot was utilized to examine the cleavage of poly (ADP-ribose) polymerase (PARP), a marker of apoptosis. RESULTS: A dose-dependent increase in phosphorylation of GSK-3beta was observed after treatment with both ZM336372 and LiCl. Growth inhibition due to treatment with ZM336372 and LiCl also occurred in a dose-dependent fashion. An increase in cleaved PARP was demonstrated after treatment with both agents, as was seen previously with GSK-3beta inhibition in pancreatic adenocarcinoma cells. CONCLUSION: This is the first description of growth inhibition and apoptosis in pancreatic cancer cells related to GSK-3beta inhibition through treatment with ZM336372.

[A German network for regional anaesthesia of the scientific working group regional anaesthesia within DGAI and BDA]. / Das Netzwerk Regionalanästhesie des wissenschaftlichen Arbeitskreises Regionalanästhesie der DGAI und des BDA.

Regional anaesthesia generally is considered to be safe. However, reports of complications with different severities are also well known. The scientific working group of regional anaesthesia of the DGAI has founded a network in conjunction with the BDA. With the aid of a registry, we are now able to describe risk profiles and associations in case of a complication. Moreover, a benchmark has been implemented in order to continuously improve complication rates.

Large volume autologous plasma predonation and retransfusion in orthopedic surgical patients.

BACKGROUND: Intraoperative washed autologous transfusion of the scavenged blood can reduce the deterioration of anemia, even during the operation with a comparatively large blood loss. On the other hand, plasma level can not be collected by this system. The preoperative donation and perioperative retransfusion of autologous plasma may reduce the plasma dilution. PURPOSE: The influence of a large volume plasma predonation and perioperative retransfusion on the plasma protein level was investigated. METHODS: Thirteen patients (63.2 +/- 13.2 yr, 70.3 +/- 12.1 kg) were examined regarding their serum protein (SP), IgG, coagulation systems, colloid osmotic pressure (COP), blood cell count before, just after, 2 h after and 7 days after the donation of 900 ml plasma by plasmapheresis with a simultaneous volume replacement. Twenty surgical patients (52.8 +/- 17.3 yr, 72.6 +/- 16.6 kg, the mean predonated autologous plasma: 2100 ml) with intra- and postoperative retransfusion of autologous plasma were examined perioperatively for SP, IgG, coagulation systems and COP. These parameters were compared with that of the predonated plasma. RESULTS: All data including SP, coagulation and COP, with the exception of IgG, completely recovered within 7 days after preoperative plasmapheresis. Perioperatively, autologous washed blood transfusion system was used. The retransfused volume of autologous predonated plasma was 1740 ml on average. Although about 41 of blood on average was lost perioperatively, only one patient out of 20 patients had to be administered homologous red blood cell transfusion. The levels of most parameters, except for COP, constantly recovered in accordance with the autologous plasma transfusion. Differences in the patterns of improvement were also observed between the parameters. CONCLUSION: A 900 ml plasma predonation can therefore be safely performed with an interval of not less than a week between the last donation and the operation. Autologous plasma retransfusion is thus considered to improve the protein levels.

A phase I pharmacodynamic trial of bortezomib in combination with doxorubicin in patients with advanced cancer.

PURPOSE: This phase I trial sought to define the toxicity, maximally tolerated dose (MTD) and pharmacodynamics of a combination of bortezomib and doxorubicin in patients with advanced malignancies. PATIENTS AND METHODS: Twenty-six patients were treated with bortezomib intravenously on days 1, 4, 8 and 11, with doxorubicin also administered intravenously on days 1 and 8, both in a 21-day cycle. Dosing ranged from 1.0 mg/m(2) of bortezomib with 15 mg/m(2) of doxorubicin to 1.5 mg/m(2) of bortezomib with 20 mg/m(2) of doxorubicin. Pharmacodynamic studies performed included assessment of levels of 20S proteasome activity and ubiquitin-protein conjugates. RESULTS: The combination of bortezomib and doxorubicin was generally well tolerated. There were two dose limiting toxicities (DLT) at dose cohort 3 (1.3 mg/m(2) bortezomib, 20 mg/m(2) doxorubicin) and 2 DLT at dose cohort 3a (1.5 mg/m(2) bortezomib, 15 mg/m(2) doxorubicin). DLT seen included neutropenia, thrombocytopenia, and neuropathy. In addition, one patient developed grade 3 central nervous system toxicity in cycle 2 (not a DLT). One patient with hormone refractory prostate cancer had a partial response. Proteasome inhibition in whole blood was demonstrated and an increase in ubiquitin-protein conjugates was observed in peripheral blood mononuclear cells of most patients. CONCLUSIONS: Bortezomib and doxorubicin can be administered safely. The recommended phase II dose for this 21-day cycle is bortezomib 1.3 mg/m(2 )intravenously on days 1, 4, 8 and 11, and doxorubicin 20 mg/m(2) intravenously on days 1 and 8. This combination may be of special interest in multiple myeloma, given the activity of both drugs in that disease.

ZM336372, a Raf-1 activator, causes suppression of proliferation in a human hepatocellular carcinoma cell line.

Hepatocellular carcinoma has been described to exhibit characteristics similar to that of neuroendocrine tumors (NETs). This includes similar anti-neoplastic responses to extracellular signal-regulated kinase (ERK) activation. NET cells and HepG2 cells have both shown growth inhibition with ERK activation. ZM336372, a Raf-1 activating agent, has been shown to cause growth inhibition and suppression of hormone secretion in a neuroendocrine cell line. Here we examine treatment of the HepG2 cell line with ZM336732 to determine if a similar anti-proliferative response will be obtained. HepG2 cells were treated with ZM336372 or solvent (dimethyl sulfoxide). The resulting effect on the proliferation was measured using the 3,4-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Western blot analysis was performed to examine the activation of the Raf-1/mitogen-activated protein kinase kinase/ERK pathway, chromogranin A production, and p21CIP1 level. Growth inhibition was observed with ZM336372 in a dose-dependent fashion. Minimal baseline phosphorylation of ERK 1/2 was observed; however, activation was observed after treatment with ZM336372. Chromogranin A secretion was suppressed due to treatment with ZM336372. A dose-dependent up-regulation of p21CIP1 was observed in response to ZM336372 treatment. ZM336372 causes growth inhibition, suppression of hormone secretion, and up-regulation of cell cycle inhibitors in a human hepatocellular carcinoma cell line, similar to that previously seen in NETs.

Electron paramagnetic resonance study of peripheral blood mononuclear cells from patients with refractory solid tumors treated with Triapine.

The metal chelator Triapine, 3-aminopyridine-2-carboxaldehyde thiosemicarbazone, is a potent inhibitor of ribonucleotide reductase. EPR spectra consistent with signals from Fe-transferrin, heme, and low-spin iron or cupric ion were observed in peripheral blood mononuclear cells (PBMCs) obtained from patients treated with Triapine. One signal that is unequivocally identified is the signal for Fe-transferrin. It is hypothesized that Fe uptake is blocked by reactive oxygen species generated by FeT(2) or CuT that damage transferrin or transferrin receptor. A potential source for the increase in the heme signal is cytochrome c released from the mitochondria. These results provide valuable insight into the in vivo mechanism of action of Triapine.

Posterior atlantoaxial three-point fixation: comparison of intraoperative performance between open and percutaneous techniques.

INTRODUCTION: Atlantoaxial instabilities, which require surgical fixation follow a variety of clinical disorders. Different surgical procedures are used for stabilization of the atlantoaxial complex, mainly posterior wiring techniques and transarticular screw fixation. Nowadays, often a combination of transarticular screws and a posterior one-point fixation is used to achieve a three-point fixation, with superior biomechanical stability and good clinical results. Different modifications were developed to improve this technique. In 1995, a percutaneous approach for atlantoaxial stabilization was introduced. In clinical studies, the technique showed a tendency towards better outcome. Beside the outcome, the intraoperative performance is of special interest for minimal invasive approaches. We therefore compared the operation time, screw angulation and blood loss, between the open and percutaneous posterior atlantoaxial techniques. MATERIALS AND METHODS: Two groups, each consisting of 17 patients, with either open (group 1) or percutaneous (group 2) atlantoxial stabilization, were compared. The operation time was retrospectively acquired from the patient's charts. The data for blood loss was provided by our anaesthesiological department, separated for intraoperative, postoperative and total blood loss. Screw angulation was measured on the postoperative x-ray by an orthopaedic surgeon. RESULTS: The percutaneous group showed an average intraoperative blood loss of 239.7 ml, compared to 929.4 ml for the open group (p< or =0.001). The analogue values for the postoperative blood loss were 142.9 ml and 379.4 ml for group 2 and group 1, respectively (p=0.008). Consecutively, the total blood loss showed also a statistically significant difference (p< or =0.001). The operation time was significantly different (p< or =0.001), with average values of 175.3 min (group 1) and 110.6 min (group 2). Screw angulation showed a trend towards a steeper angulation in the percutaneous group with an average angle of 56.8 degrees , compared to 53.9 degrees (group 1), although this was not statistically significant (p=0.053). CONCLUSION: The percutaneous technique for atlantoaxial stabilization with a three-point fixation has clear intraoperative benefits, with shorter operation time and reduced blood loss. A trend towards steeper screw angulation was found and shows at least equal feasibility for transarticular screw placement with the percutaneous technique, compared to the standard open approach.

Efficacy and cost effectiveness of harmonic scalpel compared with electrocautery in posterior instrumentation of the spine.

Different methods to reduce blood loss during spinal surgery have been described already. Although the use of the harmonic scalpel (HS), an ultrasonically activated coagulator, has been described in endoscopic spinal surgery, its efficacy in posterior instrumentation of the spine remains unclear. The aim of this study was to determine if blood loss was lower using the HS than electrocauterization (EC) and to evaluate the cost effectiveness of the HS in reducing the need for transfusion in patients undergoing posterior instrumentation of the spine. The two groups were matched in a blinded manner, without knowledge of blood loss and were similar with respect to mean age, diagnosis and operation data. All instrumentations were done by the same surgeon. After matching was completed (HS group n = 50, EC group n = 50) blood loss and overall costs for blood products were analyzed by independent observers. The following were significantly lower with the HS than with EC: (1) blood loss (1106+/-985 ml vs 2176+/-1764 ml, P < 0.001), (2) frequency of cell saver use (13 vs 28 patients, P = 0.001), (3) average cost of blood products (Euro 72 vs Euro 219, P < 0.001), (4) predonation of autologous fresh frozen plasma (2.58+/-2.78 vs 4.5+/-2.2 U, P = 0.002) and red blood cells (0.38+/-0.75 vs 0.88+/-1.1 U, P = 0.009). The overall costs, including the costs for the HS, remained neutral. The use of the HS in posterior spinal surgery leads to significantly lower blood loss, and less need for and cost of blood products, compared to EC in cases with major anticipated blood loss.

Phase I pharmacokinetic and pharmacodynamic study of recombinant human endostatin in patients with advanced solid tumors.

PURPOSE: Endostatin is the first endogenous angiogenesis inhibitor to enter clinical trials. Laboratory investigations with endostatin have indicated broad antitumor activity coupled with remarkably low toxicity. A phase I trial of recombinant human endostatin was designed to evaluate toxicity and explore biologic effectiveness in patients with refractory solid tumors. PATIENTS AND METHODS: Endostatin was administered as a 1-hour intravenous infusion given daily for a 28-day cycle. A starting dose of 30 mg/m2 was explored with subsequent dose escalations of 60, 100, 150, 225, and 300 mg/m2. Assessment of serum pharmacokinetics was performed on all 21 patients. Western blot assay and mass spectroscopy were employed to evaluate endostatin metabolism. Circulating levels of endogenous proangiogenic growth factors were examined. Tumor and tumor blood supply were imaged by dynamic computed tomography (CT), magnetic resonance imaging, ultrasound, and positron emission tomography. RESULTS: Endostatin given on this schedule was essentially free of significant drug-related toxicity. Two transient episodes of grade 1 rash were observed. No clinical responses were observed. Endostatin pharmacokinetics were linear with dose, and serum concentrations were achieved that are associated with antitumor activity in preclinical models. No aggregate effect on circulating proangiogenic growth factors were seen, although several patients exhibited persistent declines in vascular endothelial growth factor levels while enrolled in the study. A few patients demonstrated changes in their dynamic CT scans suggestive of a decline in microvessel density, although overall, no consistent effect of endostatin on tumor vasculature was seen. CONCLUSION: Endostatin given daily as a 1-hour intravenous infusion was well tolerated without dose-limiting toxicity at doses up to 300 mg/m2.