Comparison of 177Lu-octreotate and 177Lu-octreotide for treatment in human neuroblastoma-bearing mice.

Background: Patients with high-risk neuroblastoma (NB) have a 5-year event-free survival of less than 50 %, and novel and improved treatment options are needed. Radiolabeled somatostatin analogs (SSTAs) could be a treatment option. The aims of this work were to compare the biodistribution and the therapeutic effects of 177Lu-octreotate and 177Lu-octreotide in mice bearing the human CLB-BAR NB cell line, and to evaluate their regulatory effects on apoptosis-related genes. Methods: The biodistribution of 177Lu-octreotide in mice bearing CLB-BAR tumors was studied at 1, 24, and 168 h after administration, and the absorbed dose was estimated to tumor and normal tissues. Further, animals were administered different amounts of 177Lu-octreotate or 177Lu-octreotide. Tumor volume was measured over time and compared to a control group given saline. RNA was extracted from tumors, and the expression of 84 selected genes involved in apoptosis was quantified with qPCR. Results: The activity concentration was generally lower in most tissues for 177Lu-octreotide compared to 177Lu-octreotate. Mean absorbed dose per administered activity to tumor after injection of 1.5 MBq and 15 MBq was 0.74 and 0.03 Gy/MBq for 177Lu-octreotide and 2.9 and 0.45 Gy/MBq for 177Lu-octreotate, respectively. 177Lu-octreotide treatment resulted in statistically significant differences compared to controls. Fractionated administration led to a higher survival fraction than after a single administration. The pro-apoptotic genes TNSFS8, TNSFS10, and TRADD were regulated after administration with 177Lu-octreotate. Treatment with 177Lu-octreotide yielded regulation of the pro-apoptotic genes CASP5 and TRADD, and of the anti-apoptotic gene IL10 as well as the apoptosis-related gene TNF. Conclusion: 177Lu-octreotide gave somewhat better anti-tumor effects than 177Lu-octreotate. The similar effect observed in the treated groups with 177Lu-octreotate suggests saturation of the somatostatin receptors. Pronounced anti-tumor effects following fractionated administration merited receptor saturation as an explanation. The gene expression analyses suggest apoptosis activation through the extrinsic pathway for both radiopharmaceuticals.

James F. Lind Sr. MD, CM, FACS, FRCC: a tribute to a surgical gastroenterologist and educator.

BACKGROUND: This manuscript is a tribute to James Forest Lind Sr., who died on October 22, 2010. He was a founding past president of the Society of American Gastrointestinal Surgeons and devoted his life to surgical gastroenterology and academic surgery. METHODS: This manuscript was prepared by reviewing his life with input from family, friends, and colleagues. It represents a compilation of a 50-year career in medicine with enormous contributions to gastrointestinal surgery and education. RESULTS: Jim Lind Sr. was born in Fillmore, Saskatchewan, Canada on November 22, 1925. He spent his formative years in Keewatin, Ontario, Canada. After graduating from high school, he served in the Royal Canadian Air Force and then received his MD degree from Queens University. He went on to surgical internship at Hamilton General Hospital, followed by a surgical residency at Queens University. Following residency, he spent 2 years in England as a senior registrar and then completed a fellowship in gastrointestinal physiology at the Mayo Clinic. Jim Lind took his first academic appointment when he joined the faculty at the University of Manitoba in Winnipeg, Manitoba. During his distinguished career in academic surgery, Jim Lind served as the chairman of surgery at three intuitions: the University of Manitoba, McMaster University, and Eastern Virginia Medical School. CONCLUSIONS: James F. Lind Sr.'s legacy as a human being will be that he always pursued excellence in everything he did. He lived and breathed academic surgery and was passionate about surgical education. He will be missed by family, friends, and colleagues.

A portable palpation training platform with virtual human patient.

Palpation (the application of touch to the surface of the body) is an essential clinical skill. Correct palpation is part of a complete physical examination and it assists a clinician in making an accurate diagnosis, while poor palpatory skills can lead to diagnostic errors. As with any clinical skill, palpation is best learned through repetitive practice with constructive feedback. Unfortunately, changes in healthcare provide fewer opportunities for hands-on learning of this essential skill. Unlike other clinical skills, palpation has no immediate feedback to the learner regarding their performance. For example, when students are learning how to insert an intravenous catheter, failure to perform the technique correctly results in no blood return in the catheter. However, students do not know if they are palpating an abnormality if they have never felt it before. This inherent difficulty makes expert feedback even more vital to learning correct palpation. Existing research tools have addressed some of these challenges through simulation techniques that do not require experts, and can provide feedback on palpation pressures and palpation patterns. We describe a novel computer-based palpation training system, leveraging existing approaches, with an emphasis on sensing accuracy, directed-feedback, portability, and user experience.

Mixed reality humans: evaluating behavior, usability, and acceptability.

This paper presents Mixed Reality Humans (MRHs), a new type of embodied agent enabling touch-driven communication. Affording touch between human and agent allows MRHs to simulate interpersonal scenarios in which touch is crucial. Two studies provide initial evaluation of user behavior with a MRH patient and the usability and acceptability of a MRH patient for practice and evaluation of medical students' clinical skills. In Study I (n=8) it was observed that students treated MRHs as social actors more than students in prior interactions with virtual human patients (n=27), and used interpersonal touch to comfort and reassure the MRH patient similarly to prior interactions with human patients (n=76). In the within-subjects Study II (n=11), medical students performed a clinical breast exam on each of a MRH and human patient. Participants performed equivalent exams with the MRH and human patients, demonstrating the usability of MRHs to evaluate students' exam skills. The acceptability of the MRH patient for practicing exam skills was high as students rated the experience as believable and educationally beneficial. Acceptability was improved from Study I to Study II due to an increase in the MRH's visual realism, demonstrating that visual realism is critical for simulation of specific interpersonal scenarios.

Use of a hybrid-abdominal wound simulated patient in the ACS/ASE medical student simulation skills curriculum.

OBJECTIVE: We incorporated a hybrid-abdominal wound simulation to teach/assess the acquisition of three essential clinical skills in the ACS/ASE Medical Student Simulation-based Surgical Skills Curriculum. METHOD: Third year students (N = 43) attended a workshop based on the ACS/ASE surgical skills curriculum for drain care/removal, staple removal and Steri-Strip application. Following a didactic session and demonstration using a simulated patient, student skill acquisition was assessed using the ACS/ASE module rating tool. Student interest/perceived usefulness of the workshop was evaluated using Keller's Motivational Survey. RESULTS: We used median scores to identify low proficiency (n = 20; scores 17-28) and high proficiency (n = 23; scores 29-35) groups. The high proficiency group was more knowledgeable, performed better drain care, had a higher global score and was more confident than the low proficiency group. The students rated the workshop highly based on the Keller's Motivational Survey. CONCLUSION: All students were proficient in the procedure tasks and communication skills and most felt that the course was beneficial. The ACS/ASE Medical Student Simulation-based Surgical Skills Curriculum was successfully integrated into our third year surgical clerkship.

Medical student satisfaction using a virtual patient system to learn history-taking communication skills.

Virtual patients (VPs) have the potential to augment existing medical school curricula to teach history-taking and communication skills. A goal of our current efforts to study virtual characters in health professions education is to develop a system that can be independently accessed and thus user satisfaction is an important factor in how readily this technology will be adopted. Twenty-three medical students participated in a study in which they interviewed a virtual patient and were asked to rate the educational value of the experience. Despite some of the limitations in this developing technology, students were generally receptive to its use as an educational tool. Further enhancements to the system, including increased fidelity of the interaction and novel feedback mechanisms, should improve learner satisfaction with and adoption of the virtual patient system.

New Trends at 25 Years: An American Board of Surgery Academic Certifying Examination Review Course.

BACKGROUND: In the 1980s, a small research group began identifying variables affecting applicant success on the American Board of Surgery (ABS) Certifying Examination (CE). We now report success and trends as we complete 25 years. We had multiple challenges as identified through faculty focus groups and participant feedback that needed to be addressed: increase the national optics of the program, integrate new innovative experiences, maintain the integrity of the collected data on excel files, incorporate national trends in surgery, attract experienced clinical volunteer faculty and staff, security of capital, and schedule management. METHOD: The primary purpose of the program is to define the root cause interfering with success on the ABS CE. All of the listed changes in course design (2012-2016) were entered into excel files along with participants demographics, including results of the pretesting modules, the communication inventory, all self-reported stressors, and interview results to track the effect of faculty interventions, trends and ABS outcomes. RESULTS: The profiles of the participants have changed over time, including: marital status, presence of DSM-5 stressors, gender, fellowship training, study habits, financial burdens, and international graduate status. International graduates demonstrated communication issues that were present, though rarely addressed, during residency training. The gradual absorption of junior faculty allowed a seamless transition over time as part of the succession plan. Although the national success rates on the CE were 72% to 80%, this program's success rate still remained in the 90 percentile (94%-97%) for those who followed their education improvement plan. Deidentified excel files will be converted to REDCap for preservation and analysis. DISCUSSION: The small course design has continued to be effective at identifying variables that interfere with success on the CE examination. The inclusion of additional PhD education scientists facilitated focused individual interventions. A pilot program for international graduate status residents is in development.

How Safe Is the Safety Net? Comparison of Ivor-Lewis Esophagectomy at a Safety-Net Hospital Using the NSQIP Database.

BACKGROUND: Recent data suggest that surgical outcomes at hospitals caring for low-income, vulnerable populations are suboptimal compared with outcomes from nonsafety-net hospitals. Therefore, the purpose of our study was to compare outcomes for patients who underwent an Ivor-Lewis esophagectomy at a safety-net hospital with the National Surgical Quality Improvement Program (NSQIP) database. STUDY DESIGN: We retrospectively reviewed the medical records of consecutive patients who underwent an Ivor-Lewis esophagectomy, between September 2013 and January 2017, at a single safety-net hospital. Patient characteristics and outcomes were compared with the 2013 to 2015 NSQIP database. Continuous variables were compared using Student's t-test, and categorical variables were analyzed using chi-square tests. Values of p < 0.05 were considered significant. RESULTS: We identified 78 patients from the safety-net hospital and 1,825 patients in the NSQIP database who underwent an Ivor-Lewis esophagectomy. Baseline characteristics were similar, except the safety-net hospital patients were more likely to have COPD (19.2% vs 8.1%; p = 0.001) and be current smokers (42.3% vs 26.0%; p = 0.001); patients in the NSQIP group had a higher BMI (28 kg/m2 vs 26 kg/m2; p = 0.001). There were no differences between groups for mortality, readmission, discharge destination, or mean operative time. Safety-net hospital patients had significantly fewer complications (16.7% vs 33.3%; p = 0.003), fewer reoperations (6.4% vs 14.5%; p = 0.046), and shorter hospital length of stay (10.3 vs 13.1 days; p = 0.001). CONCLUSIONS: Patients who underwent an Ivor-Lewis esophagectomy at a safety-net hospital had fewer complications and reoperations, and a shorter hospital length of stay compared with a national cohort. These findings illustrate the value of clinical pathways in optimizing the patient outcomes at safety-net hospitals and providing excellent care to their vulnerable patient population.

Survey of student education in surgery.

BACKGROUND: Few data exist comparing medical student surgical education programs across the US and Canada. We conducted a 30-question, Web-based survey of 142 medical schools in the US and Canada about the administration of undergraduate surgical education programs at their institutions. STUDY DESIGN: From October 2005 to January 2006, 142 Association of American Medical Colleges (AAMC) medical schools in the US and Canada were asked to complete a 30-item, Web-based questionnaire. Descriptive statistics were compiled for all responses and data were analyzed using Student's t-test. RESULTS: One hundred two medical schools responded to the survey (90 of 125 US and 12 of 17 Canadian schools, overall response rate,=72%). The majority of surgical clerkships have between 16 and 30 students rotating at a time through 2 to 4 teaching sites. The most frequent overall clerkship duration was 8 weeks, divided most frequently between a 4- to 6-week general surgery rotation and a 4-week subspecialty rotation. There was no consensus about what services constitute general surgery rotations, although the most common subspecialty rotations were urology (68%) and orthopaedics (66%). American medical schools were more likely to use the National Board of Medical Examiners (NBME) subject examination for student assessment (90.9% US versus 50.0% Canadian, p < 0.05); Canadian medical schools were more likely to use an Objective Structured Clinical Examination (69.2% Canadian versus 37.7% US, p < 0.05) and other written examinations (69.2% Canadian versus 37.7% US, p < 0.05). CONCLUSIONS: There is wide variation in the structure and administration of medical student surgical education programs in the US and Canada. These findings underscore the need for a clerkship directors committee and a national dialog about a core student curriculum to ensure consistency in the quality of student education in surgery.

Comparing interpersonal interactions with a virtual human to those with a real human.

This paper provides key insights into the construction and evaluation of interpersonal simulators--systems that enable interpersonal interaction with virtual humans. Using an interpersonal simulator, two studies were conducted that compare interactions with a virtual human to interactions with a similar real human. The specific interpersonal scenario employed was that of a medical interview. Medical students interacted with either a virtual human simulating appendicitis or a real human pretending to have the same symptoms. In Study I (n = 24), medical students elicited the same information from the virtual and real human, indicating that the content of the virtual and real interactions were similar. However, participants appeared less engaged and insincere with the virtual human. These behavioral differences likely stemmed from the virtual human's limited expressive behavior. Study II (n = 58) explored participant behavior using new measures. Nonverbal behavior appeared to communicate lower interest and a poorer attitude toward the virtual human. Some subjective measures of participant behavior yielded contradictory results, highlighting the need for objective, physically-based measures in future studies.

Acquisition, retention and transfer of simulated laparoscopic tasks using fNIR and a contextual interference paradigm.

BACKGROUND: Using functional near infrared spectroscopy, a noninvasive, optical brain imaging tool that monitors changes in hemodynamics within the prefrontal cortex (PFC), we assessed performance and cognitive effort during the acquisition, retention and transfer of multiple simulated laparoscopic tasks by novice learners within a contextual interference paradigm. METHODS: Third-year medical students (n = 10) were randomized to either a blocked or random practice schedule. Across 3 days, students performed 108 acquisition trials of 3 laparoscopic tasks on the LapSim® simulator followed by delayed retention and transfer tests. Performance metrics (Global score, Total time) and hemodynamic responses (total hemoglobin (µm)) were assessed during skill acquisition, retention and transfer. RESULTS: All acquisition tasks resulted in significant practice schedule X trial block interactions for the left medial anterior PFC. During retention and transfer, random performed the skills in less time and had lower total hemoglobin change in the right dorsolateral PFC than blocked. CONCLUSIONS: Compared with blocked, random practice resulted in enhanced learning through better performance and less cognitive load for retention and transfer of simulated laparoscopic tasks.

The use of virtual patients to teach medical students history taking and communication skills.

BACKGROUND: At most institutions, medical students learn communication skills through the use of standardized patients (SPs), but SPs are time and resource expensive. Virtual patients (VPs) may offer several advantages over SPs, but little data exist regarding the use of VPs in teaching communication skills. Therefore, we report our initial efforts to create an interactive virtual clinical scenario of a patient with acute abdominal pain to teach medical students history-taking and communication skills. METHODS: In the virtual scenario, a life-sized VP is projected on the wall of an examination room. Before the virtual encounter, the student reviews patient information on a handheld tablet personal computer, and they are directed to take a history and develop a differential diagnosis. The virtual system includes 2 networked personal computers (PCs), 1 data projector, 2 USB2 Web cameras to track the user's head and hand movement, a tablet PC, and a microphone. The VP is programmed with specific answers and gestures in response to questions asked by students. The VP responses to student questions were developed by reviewing videotapes of students' performances with real SPs. After obtaining informed consent, 20 students underwent voice recognition training followed by a videotaped VP encounter. Immediately after the virtual scenario, students completed a technology and SP questionnaire (Maastricht Simulated Patient Assessment). RESULTS: All participants had prior experience with real SPs. Initially, the VP correctly recognized approximately 60% of the student's questions, and improving the script depth and variability of the VP responses enhanced most incorrect voice recognition. Student comments were favorable particularly related to feedback provided by the virtual instructor. The overall student rating of the virtual experience was 6.47 +/- 1.63 (1 = lowest, 10 = highest) for version 1.0 and 7.22 +/- 1.76 for version 2.0 (4 months later) reflecting enhanced voice recognition and other technological improvements. These overall ratings compare favorably to a 7.47 +/- 1.16 student rating for real SPs. CONCLUSIONS: Despite current technological limitations, virtual clinical scenarios could provide students a controllable, secure, and safe learning environment with the opportunity for extensive repetitive practice with feedback without consequence to a real or SP.

Virtual patients: assessment of synthesized versus recorded speech.

Virtual patients have great potential for training patient-doctor communication skills. There are two approaches to producing the virtual human speech: synthesized speech or recorded speech. The tradeoffs in flexibility, fidelity, and cost raise an interesting development decision: which speech approach is most appropriate for virtual patients? Two groups of medical students participated in a user study interviewing a virtual patient under each condition. We found no significant differences in the overall impression, speech intelligibility, and task performance. Our conclusion is that if the goal is to train students of which questions to ask, synthesized speech is just as effective as recorded speech. However, if the goal is to teach the student how to ask the correct questions, a high level of expressiveness in the virtual patient is needed. This in turn necessitates the higher cost - even with the lower flexibility - of recorded speech.

Minimizing local recurrence after breast conserving therapy using intraoperative shaved margins to determine pathologic tumor clearance.

BACKGROUND: This study analyzed survival, locoregional recurrence, and reexcision rates after breast conserving therapy, based on the margin analysis technique used at the University of Florida, which incorporates frozen section analysis of shaved breast tissue from the lumpectomy cavity. STUDY DESIGN: A retrospective review was done of 257 patients who underwent 267 consecutive lumpectomy operations and completed radiation therapy at our institution. Margins in 189 patients were assessed using frozen section analysis of shaved margins from the lumpectomy cavity. RESULTS: Breast conserving therapy was performed for 220 (83%) patients with early breast cancer (T1 and T2 tumors) and 47 (17%) with ductal carcinoma in situ. With a median followup of 5.6 years, the crude locoregional recurrence rates for patients who had margins analyzed intraoperatively by frozen section analysis or margins analyzed by permanent analysis were 1.9% (3 of 157) and 3.1% (2 of 63), respectively, for early breast cancer and 15.6% (5 of 32) and 6.6% (1 of 15) for ductal carcinoma in situ (p=NS). Survival rates were 97% and 78%, at 5 and 10 years, respectively, for the early breast cancer patients, and 98% and 98%, respectively, for ductal carcinoma in situ patients. Permanent intraoperative frozen section analysis reexcision rates were 33.3% (26 of 78) and 5.8% (11 of 189). CONCLUSIONS: Regardless of the technique used for margin analysis, breast conserving therapy led to low locoregional recurrence relative to national figures, pointing to the importance of the technique of radiation therapy at our institution. Reexcision rate was reduced with the use of frozen section analysis.

External beam radiotherapy for primary and adjuvant management of aggressive fibromatosis.

PURPOSE: To review a large single-institution experience in the management of aggressive fibromatosis to determine the effectiveness of external beam radiotherapy (EBRT) and identify the presentation and treatment variables predictive of locoregional control. METHODS AND MATERIALS: Between 1975 and 2000, 72 patients were treated with EBRT for a pathologically confirmed diagnosis of aggressive fibromatosis. Thirty patients were treated at the primary presentation and 42 at the time of a locoregional recurrence. Minimal 2-year follow-up data were available for 65 patients (median 6 years). Megavoltage irradiation with 60Co to 20 MV photons or electron therapy was used for all patients. Most patients were treated after attempted complete surgical resection; 16 patients underwent pretreatment biopsy alone. The prescribed treatment was standard (1.8 Gy) daily fractions in 42 cases and 1.2 Gy fractions b.i.d. in 23 cases. The median prescribed dose was 54 Gy. The prognostic variables and treatment results were evaluated by Kaplan-Meier actuarial analysis. RESULTS: Locoregional control was achieved in 52 of 65 patients. The 5-year actuarial locoregional control was 83%. Locoregional failure occurred in 13 patients (11 in patients with recurrent tumors). Only two failures occurred within the irradiation fields; nine failures occurred at the field margins. Eleven patients were salvaged by surgery: wide excision in nine and amputation in two. The only prognostic factor significant for locoregional control was primary vs. recurrent presentation (p = 0.0193). The 5-year locoregional control rates for irradiation at initial presentation and at recurrence were 96% and 75%, respectively. The variables without significance for locoregional control included primary tumor location, surgical procedures performed, resection margins, and gross vs. microscopic residual tumor at irradiation. Lymphedema was the most common late effect, occurring in 7 patients, 5 with prior treatment. Bone fracture occurred in 3 patients; all 3 had fibromatosis involving the bone at presentation but without recurrence at the time of fracture. CONCLUSION: EBRT is effective treatment for aggressive fibromatosis. The probability of locoregional control decreases with multiple prior recurrences.

Prevalence of fra(X) in the county of Funen in Denmark is lower than expected.

Population based cytogenetic fra(X) surveys have not previously been reported from Denmark. In the present study we present an estimate of fra(X) based on 1) information from the Danish Central Cytogenetic Registry of diagnosed fra(X) males of all age groups in all Denmark in the period 1980-1992 and 2) a systematic cytogenetic fra(X) survey of 175 of 8-10-year-old children with special educational needs resident in a defined, demographically representative area of Denmark (the county of Funen). The study was performed in 1988-90 before the cloning of the FMR-1 gene. In the county of Funen there were 7,837 male children in the age group of 8-10 years. In the cytogenetic survey of learning disabled children, no fra(X) positive was diagnosed. There were 99 registered males with fra(X) in all Denmark, equivalent to a prevalence of 0.04 per 1,000 males (confidence interval 0.032:1,000-0.048:1,000). Molecular fra(X) surveys of different, large populations are needed in order to estimate the frequency of fra(X) and clarify whether significant differences in prevalence exist in different populations.

Endotoxin stimulates arginine transport in pulmonary artery endothelial cells.

BACKGROUND: The pulmonary endothelium plays an important role in the metabolism of the amino acid arginine, the exclusive precursor molecule for nitric oxide (NO). Despite decreased circulating arginine levels, endothelial NO production is elevated during endotoxemia. However, the regulation of pulmonary artery endothelial arginine transport has not been studied. We hypothesized that endotoxin stimulates carrier-mediated arginine transport by the pulmonary endothelium. METHODS: The relative contributions of the various transport systems to total arginine transport by porcine pulmonary artery endothelial cells (PAECs) was determined by assaying the uptake of 3H-L-arginine in the presence or absence of Na+. PAECs were then incubated with various concentrations of Escherichia coli endotoxin, and y(+)-mediated arginine transport was measured at different time points thereafter. Kinetic studies were performed over a range of arginine concentrations to determine changes in transport affinity and maximum rate of metabolism. To address the role of RNA and protein synthesis in the increased transport, uptake was measured after exposure of cells to the transcriptional inhibitor actinomycin D and the protein synthesis inhibitor cycloheximide. RESULTS: Most (75%) of arginine transport by PAECs was mediated by the high-affinity Na(+)-independent transport system y+. Endotoxin stimulated y(+)-mediated arginine transport by PAECs twofold to fivefold, a response that was time and dose dependent. The accelerated transport was detectable within 2 hours and maximal at 12 hours. Kinetic studies revealed that the accelerated arginine transport was the result of a 68% increase in the maximal transport velocity (1519 +/- 65 pmol/mg protein/30 sec in endotoxin-treated cells vs 903 +/- 96 in control cells; p < 0.01) without a change in transport affinity. The endotoxin-mediated increase in arginine uptake was abrogated by actinomycin D and cycloheximide. CONCLUSIONS: Endotoxin stimulates Na(+)-independent arginine transport by PAECs through a process that requires de novo RNA and protein synthesis, possibly of the transporter itself. This response may be designed to support arginine-dependent biosynthetic pathways in the lung during septic states.

Cytokines regulate endotoxin stimulation of endothelial cell arginine transport.

BACKGROUND: Endotoxin (lipopolysaccharide) stimulates transmembrane L-arginine transport in pulmonary artery endothelial cells (PAECs). The proinflammatory cytokines tumor necrosis factor (TNF) and interleukin-1 (IL-1) mediate many of the pathophysiologic effects of endotoxemia and sepsis. Endothelial cells secrete TNF and IL-1 in response to endotoxin. We hypothesize that lipopolysaccharide stimulation of plasma membrane L-arginine transport is mediated via an autocrine cytokine loop involving TNF and IL-1. METHODS: Confluent porcine PAECs were incubated with various concentrations of lipopolysaccharide, TNF, or IL-1, and arginine uptake was determined by assaying the uptake of 3H-L-arginine in the presence or absence of Na+ at different time points. PAECs were then incubated with lipopolysaccharide or saline solution after pretreatment with either anti-TNF antibody or IL-1-receptor antagonist, and transport was measured 12 hours later. RESULTS: Lipopolysaccharide, IL-1, and TNF all increased both Na+-dependent and Na+-independent carrier-mediated L-arginine transport in a fashion that was both time and dose dependent. Maximal increases in stimulated arginine uptake occurred 8 hours after exposure to the cytokines and 12 hours after exposure to lipopolysaccharide. Pretreatment of endothelial cells with anti-TNF antibody blocked lipopolysaccharide stimulation of both Na+-independent and Na+-dependent transport by 100% and 90%, respectively. In addition, IL-1-receptor antagonist inhibited lipopolysaccharide stimulation of both Na+-independent and Na+-dependent transport by 65% and 85%, respectively. CONCLUSIONS: The marked increase in carrier-mediated L-arginine transport activity produced by lipopolysaccharide, IL-1, and TNF may represent an adaptive response by the pulmonary endothelium to support arginine-dependent biosynthetic pathways during sepsis. Furthermore, lipopolysaccharide stimulation of arginine transport is mediated in part through an autocrine mechanism involving IL-1 and TNF.

Nuclear factor-kappa B is upregulated in colorectal cancer.

BACKGROUND: Chemoresistance may involve the anti-apoptotic transcriptional regulator, nuclear factor-kappa B (NF-kappa B). The purpose of this study was to determine whether chemotherapy induces NF-kappa B activation in a human colon cancer cell line (SW48) and whether NF-kappa B is constitutively activated in colorectal cancer. METHODS: SW48 cells were incubated with gemcitabine hydrochloride (Gemzar) in the presence and absence of the 26s proteasome inhibitor, MG132, and NF-kappa B binding (electrophoretic mobility shift assay), DNA synthesis (tritiated thymidine uptake), cell viability (3-[4,5-dimethylthiazol-2-yl]-diphenyl-tetrazolium bromide assay), and apoptosis (caspase-3 activity) were measured at 24 hours. NF-kappa B binding (electrophoretic mobility shift assay) was also assayed in 10 colorectal cancer tumors. RESULTS: SW48 cells demonstrated constitutive NF-kappa B binding that was enhanced by gemcitabine hydrochloride in a dose-dependent manner. MG132 inhibited NF-kappa B binding and enhanced gemcitabine hydrochloride's inhibition of DNA synthesis (gemcitabine hydrochloride = 73% +/- 1.4% vs gemcitabine hydrochloride + MG132 = 6% +/- 0.4%, P <.05), cell killing (gemcitabine hydrochloride = 87% +/- 2.0 vs gemcitabine hydrochloride + MG132 = 25% +/- 1.3%, P <.05), and caspase-3 activity (gemcitabine hydrochloride = 870 +/- 17.4 vs gemcitabine hydrochloride + MG132 = 1075 +/- 20.4, P <.05). NF-kappa B binding was increased in 8 of 10 colorectal cancer tumors compared with adjacent normal mucosa. CONCLUSIONS: Gemcitabine hydrochloride enhances NF-kappa B binding in a colorectal cancer cell line, whereas inhibition of NF-kappa B enhances gemcitabine hydrochloride's antitumor activity. NF-kappa B is also activated in human colorectal cancer. NF-kappa B may identify chemoresistant tumors, whereas inhibition of NF-kappa B may be a novel, biologically based therapy. (Surgery 2001;130:363-9).

Endotoxin-induced nitric oxide production in pulmonary artery endothelial cells is regulated by cytokines.

BACKGROUND: L-Arginine is the sole precursor of nitric oxide (NO). Bacterial lipopolysaccharide (endotoxin) (LPS) stimulates carrier-mediated L-arginine transport in porcine pulmonary artery endothelial cells (PAECs) through an autocrine pathway that involves interleukin-1 alpha (IL-1 alpha) and tumor necrosis factor alpha (TNF-alpha). OBJECTIVES: To determine if Escherichia coli LPS stimulates NO synthesis in PAECs and, if so, if LPS stimulation of NO production is also mediated by autocrine secretion of IL-1 alpha and TNF-alpha. DESIGN: Monolayers of PAECs were incubated with various concentrations of LPS, recombinant human TNF-alpha, or IL-1 alpha, and total nitrate-nitrite accumulation was measured at different time points with the Greiss reagent following cadmium reduction. Release of TNF-alpha and IL-1 alpha release by LPS-stimulated PAECs were measured using the WEHI (for TNF-alpha) and A375.S2 (for IL-1 alpha) bioassays. The PAECs were then incubated with saline solution or LPS in the presence or absence of either a polyclonal antibody to human TNF or IL-1 receptor antagonist, and nitrate-nitrite accumulation was measured at 48 hours. RESULTS: Production of NO by PAECs was increased 230% by LPS (1 microgram/mL), 350% by TNF-alpha (1000 U/mL), and 240% by IL-1 alpha (1000 U/mL) (P < .05 vs control). The LPS-stimulated NO production was inhibited by IL-1 receptor antagonist (100 micrograms/mL) or antibody to TNF (10 micrograms/mL) to control levels (P < .05 vs LPS; difference vs saline solution was not significant). The LPS-stimulated TNF-alpha secretion by PAECs and TNF-alpha activity were maximal at 6 hours (400 +/- 42 pg/mL). The IL-1 alpha activity was not detectable in LPS-stimulated PAECs by the A375.S2 bioassay. CONCLUSIONS: Endotoxin, TNF-alpha, and IL-1 alpha stimulated NO synthesis in PAECs. Endotoxin-stimulated NO synthesis through an autocrine pathway involving the cytokines TNF-alpha and IL-1 alpha.