63.18 Factors Associated with Patient Delay to Surgical Presentation in Cameroon

Posted on December 22, 2014

C. Long1, T. T. Ngwa2, E. K. Lawong2, J. A. Brown2, S. M. Wren1  1Stanford University,Palo Alto, CA, USA 2Mbingo Baptist Hospital,Mbingo, NORTH-WEST, Cameroon

Introduction: Delay in definitive management for certain surgical diseases is correlated with worse outcomes and can be present in health systems regardless of country economic status. Few studies have critically examined delays in Sub-Saharan Africa. Reasons are often multi-factorial including cultural, socioeconomic, and access issues. This study investigates factors contributing to delays in treatment at Mbingo Baptist Hospital (MBH), a referral hospital in Cameroon and West Central Africa.

Methods: 235 patients and/or caregivers were interviewed (6/23/14-8/4/14) using a 34-item survey. All patients >age 15 admitted with trauma, cancer, or emergent surgical conditions (defined as requiring a definitive procedure within 48 hours of admission) were interviewed and demographics, history of current illness, previous care received, and factors that contributed to delayed arrival were collected.

Results: Delays were scored as <1 day, 1-6 days, 1-2 weeks, 2 weeks-1 month, 1-6 months, 6-12 months, 1-5 years, and 5-10 years. Of the 235 patients, 105 (44.6%) had trauma, 78 (33.2%) emergent conditions, and 52 (22.1%) cancer. Presence of symptoms were found >1 month in100% of cancer patients, >2 weeks in 78.2% of emergent patients, and > 1 week in 47.6% of trauma patients before presentation to MBH. The most common reason for seeking medical help was pain (71.8%, n=168), broken down to 67% (35/52) for cancer, 71.4% (75/105) for trauma, and 74.3% (58/78) for emergencies. The greatest factor for delay was at least one visit to prior healthcare facility in 88.9%.  By condition, it was 100% (52/52) for cancer, 85.7% (90/105) for trauma, and 85.9% (67/78) for emergencies.  A total of 65.5% (n=137) received previous care at a government hospital, 81.3% (n=170) received medication to “treat” their condition, and 16.3% (n=34) received a surgical operation. 99% (n=207) did not feel that their health concerns were addressed at that encounter. A total of 56.5% (n=118) had received care in 2 or more different places with only 10.0% (n=21) reporting that their condition improved afterwards. Other common factors identified were transportation related (46%), feeling that they were not “sick” enough (34.9%) and cost related (25.5%).

Conclusions: Significant delays in receiving definitive treatment were most often due to the time spent on previous failed attempts at care. Deciding where to seek medical help is complex, as patients and family members balanced factors such as cost, distance, severity and urgency of the condition, and the reputation of different facilities. This information can be used to inform policy discussions about optimal healthcare organization within the country.

63.19 Evaluation of Indications and Outcomes of Surgical Transfers at Three District Hospitals in Rwanda

Posted on December 22, 2014

R. G. Maine1,2,3,4, C. Habiyakare10, T. Nkurunziza5, A. Hategekimana8, P. Mizero8, W. Ndayambaje5, E. Nsengiyumva6, J. Havugimana6, J. Nigilimana9, J. Uwimungu5, J. Meara2,3, G. Ntakiyiruta6,7, J. Mubiligi5,9, R. Riviello1,2,7  1Brigham And Women’s Hospital,Center For Surgery And Public Health,Boston, MA, USA 2Harvard School Of Medicine,Program In Global Surgery And Social Change,Brookline, MA, USA 3Boston Children’s Hospital,Plastic And Oral Surgery,Boston, MA, USA 4University Of California – San Francisco,Department Of Surgery,San Francisco, CA, USA 5Inshuti Mu Buzima (Partners In Health),Kigali, , Rwanda 6University Of Rwanda,School Of Medicine,Kigali, , Rwanda 7Central Teaching Hospital Of Kigali,Department Of Surgery,Kigali, , Rwanda 8Rwinkwavu Hospital,Kayonza, , Rwanda 9Butaro Hospital,Burera, , Rwanda 10Kirehe Hospital,Kirehe, , Rwanda

Introduction:
Many countries have few surgeons and patients who need surgical care must transfer to the hospitals where those surgeons work. Few studies have evaluated the surgical transfer system in these low resource settings to understand indication for transfer and patient outcomes after transfer. This study evaluated surgical transfers at three district hospitals in Rwanda.

Methods:
We retrospectively reviewed the charts of all patients admitted to the surgical ward at these hospitals in Rwanda from January 1, 2013 to December 31, 2013. We also reviewed charts of patient who transferred from other wards to the referral center for surgical care. At the district hospitals, diagnosis, surgical treatment, and indication for transfer were collected. At the referral center treatment received and complications were collected. Complications included death, surgical site infections, pneumonia, urinary tract infection, pressure ulcers, unplanned reoperation or intubation, anastomotic leak, and other infections.

Results:

A total of 2139 surgical ward charts were reviewed at the district hospitals. An additional 299 charts of patients who transferred from other wards were reviewed. Of all surgical ward admissions, 353 (16.5%, 95%CI:14.9%-18.1%) were recommended to transfer, and 256 (72%, 95%CI:67.5%-77.1%) transferred. The percent of patient who transferred from the surgical wards at the district hospitals were 10.2%, 17.9% and 10% (p<0.001). A diagnosis was documented in 521 patients who transferred from all wards. The primary diagnoses for transferred patients include:  fractures 140 (27.3%), other trauma 76 (14.8%), cancer 67 (13.1%), and surgical infections 47 (9.2%). The indications for transfer included lack of appropriate personnel at the district hospital 82.1%, lack of equipment 27.3%, lack of supplies 2.7%, need for intensive care 5.3%, patient or family preference 1.4%,  or development of a complication at the district hospital 0.7%. The remaining 4.2% of patients transferred for an unknown reason. 

A total of 399 records were located at the referral hospital. Of the records located, 236 (59.1%, 95%CI:54.1-64%) patients received an operation at the referral hospital. This compares to 62.8%, 13.4% and 13.9% of patient admitted to the surgical ward at the three district hospitals (p <0.001). The overall complication rate for surgical ward patients at the district hospitals was 4.65: 5.7% for transferred patients and 4.4% for non-transferred patients(p =0.35). At the referral hospital, 66 patients(16.5%, 95%CI:13%-20.6%) developed a complication.

Conclusion:
Transfers for surgical care are common in district hospitals in Rwanda. The primary reason for transfer is the appropriate expertise is not available at the district hospital. Injuries are the most common diagnosis for transferred patients. Complications are common for these patients. Evaluation of skill sets of district hospital personnel and delays in surgical care for transferred patients is needed.

63.20 Utilizing Technology to Connect Educational Initiatives across the World

Posted on December 22, 2014

M. Swaroop1, E. Yang4, C. T. Richards3, M. Schuller1, S. Krishnaswami4  1Northwestern University,Trauma And Critical Care,Chicago, IL, USA 3Northwestern University,Emergency Medicine,Chicago, IL, USA 4Oregon Health And Science University,Pediatric Surgery,Portland, OR, USA

Introduction:

With the rapid increase in online collaboration and educational platforms available, clinicians and researchers in low and middle-income countries and their partners, can benefit from an introduction to these tools.  This abstract aims to describe a workshop that was designed to introduce emerging technologies that may be leveraged to strengthen cross-continent clinical, education and research collaborations.

Methods:
An Association for Academic Surgery-sponsored workshop was piloted as a parallel session at the 2013 World Congress of Surgery, Obstetrics, Trauma, and Anesthesia in October 2013 in Trinidad. During the interactive session, participants signed up for services and practiced networking using learning management systems (such as Moodle), scheduling platforms (such as Doodle), virtual meeting software (such as Oovoo or Skype), and social media tools (such as Facebook or twitter) in real-time. Potential uses for these tools in collaborative global surgery initiatives were then discussed.  Demographics of participants and evaluations from the session were analyzed.

Results:

Twenty-one of 335 congress registrants attended the Technology workshop as a parallel session during the Congress.  The majority of workshop participants (55%) were male and 70% were physicians. The remainder represented other allied health professionals or healthcare administrators.  A significant minority (25%) of participants were from low and middle income regions.  All participants who evaluated the session rated it as excellent or very good with an mean rating of 4.5 out of 5, and 90% felt the session objectives were fully met.  Themes in feedback for the workshop included its practical utility and the ability for the session to improve and enhance practice.

Conclusion:

Leveraging technology to promote educational initiatives among international collaborators on global surgery initiatives is critical to effective and timely communication.  This pilot workshop on the subject was very well-received by a small but diverse audience.   Ongoing curriculum development for such courses to match new technologies and deployment to a wide audience in both low, middle, and high income countries will be essential towards ensuring efficacy.

64.01 Anastomotic Leak After Closure of Loop Ileostomy vs. Small Bowel Resection

Posted on December 22, 2014

J. Chen1, D. T. Huynh1, C. M. Divino1  1Icahn School Of Medicine At Mount Sinai,General Surgery,New York, NY, USA

Introduction:  Loop ileostomies are often created to temporarily protect a distal anastomosis however closure of the stoma can be associated with anastomotic leak(AL) and morbidity. The aim of our study is to compare the anastomotic leak rate after loop ileostomy closure(LIC) with the leak rate after small bowel resection(SBR).

Methods:  A retrospective chart review was performed of patients at our institution from 2005-2014 for patients diagnosed with anastomotic leak after LIC and compared to those who developed an anastomotic leak after a SBR.

Results: The anastomotic leak rate after LIC was 3.16% compared to 1.16% for SBR. This difference in the leak rate between LIC and SBR was found to be statistically significant(p=0.0023). There was no significant difference found between the comorbidities of the two groups.

Conclusion: The anastomotic leak rate after LIC is 2.7 times that of the leak rate after SBR which is significantly different despite the procedures involving the same small bowel to small bowel anastomosis. We hypothesize that this is due to difficulty gaining adequate exposure through the local incision during LIC compared to SBR. 

 

64.02 Design and Experimental Evaluation of an Anti-Leak Feeding Tube

Posted on December 22, 2014

L. G. Gutwein1, R. Helmig2, L. G. Gutwein1  1Indiana University School Of Medicine,Plastic & Reconstructive Surgery,Indianapolis, IN, USA 2University Of Florida,Gainesville, FL, USA

Introduction:  Enteral feeding is commonplace in healthcare.  The present design of a gastrostomy tube is inadequate because leakage of gastric contents onto the skin is usual prompting emergency department visits and unnecessary wounding and infection that may require hospital admission or operative debridement.  It is common practice during the gastrostomy tube procedure to place a silk suture tightly around the neck portion of an external retaining member to increase the frictional force against the gastric tube, because it is widely recognized that, with time, the static friction between the tube shaft and external retaining member will inevitably be overcome by dynamic friction.  As swelling reduces and the tube materials wear, the gastrostomy balloon is able to move away from the gastric wall allowing gastric contents to leak out around the gastrostomy tube and onto the skin.  In view of the current problems, there is a need for a gastric feeding tube device that can be easily adjusted and prevent leakage by maintaining the internal retention balloon or bumper juxtaposed to the gastric mucosa. In this study, we introduce a new gastrostomy tube design and prototype that inhibits leakage by utilizing an adjustable external retaining member which compresses against the feeding tube shaft thereby preventing dynamic friction.

Methods:  A conventional external retaining member of a 22 French gastrostomy tube is tested against a novel compression fitting external retaining member.  Each gastrostomy tube was clamped to a scale and the external retaining member moved by hand to slide along the tubing at a constant rate and the applied force was recorded.

Results: An experimental prototype (Figure) was tested against the conventional design control for 10 experimental repetitions each (Group 1).  The mean forces were 18 and 46 ounces for conventional and experimental designs, respectively. The tube shaft was exchanged and the experiment repeated for 10 repetitions (Group 2).  The mean forces were 15 and 48 ounces for conventional and experimental designs, respectively.  Consistently, the experimental external retaining member demonstrated 2.5-3x the clamping frictional force as compared to the conventional design for Groups 1 and 2 (p=2.57E-13 and p=1.90E-13, respectively).  In the simulated in-situ environment with lubrication along the external shaft of the feeding tube (Group 3), the experimental external retaining member consistently performed superior to the conventional member (p=3.30E-11).

Conclusion: The refined patent-pending design has the potential to revolutionize the feeding tube market by decreasing morbidity and associated healthcare costs.

 

64.03 Laparoscopic Pedicled Omental Flap for Extraperitoneal Revascularization

Posted on December 22, 2014

M. Bruzoni1, G. Steinberg2, S. Dutta1  1Stanford University School Of Medicine,Pediatric Surgery,Stanford, CA, USA 2Stanford University School Of Medicine,Neurosurgery,Stanford, CA, USA

Introduction:  An abundance of angiogenic and immunologic factors make the omentum an ideal tissue for reconstruction and revascularization of a variety of extraperitoneal wounds and defects. Omental harvesting has historically been performed through a large laparotomy with subcutaneous tunneling to the site of disease. Complications of the open procedure include abdominal wound infection, fascial dehiscence, ventral hernia, and postoperative ileus. The use of laparoscopy to harvest the omentum has the potential to reduce such complications. We describe the outcomes of a series of patients undergoing laparoscopic pedicled omental flap mobilization for cerebral revascularization in moyamoya disease.

Methods:  A retrospective chart review of all patients undergoing laparoscopic omental-cerebral transposition for moyamoya disease between 2011 and 2014 was performed.  Clinical indication, surgical technique, operative times, complications, and outcomes at follow-up were reviewed.

Results: Seven consecutive patients ages 5 to 13 years underwent the procedure. The general surgery team performed laparoscopic omental mobilization, extraperitonealization, and subcutaneous tunneling while the neurosurgical team performed craniotomy and cerebral application of the graft. The omental flap measured up to 60 cm in length.  Operative times for the omental transposition ranged from 125 to 200 minutes.  The patients were followed up postoperatively with clinic visits and angiography.  There was one intraoperative complication (mesocolon injury requiring segmental resection in a patient with prior peritonitis) and no postoperative complications.  All patients had significant to complete symptomatic resolution, and demonstrated adequate pedicle-based revascularization on angiography. 

Conclusion: Laparoscopic pedicled omental mobilization and subcutaneous transposition is feasible and effective in children who require cerebral revascularization for moyamoya disease.  Morbidity from a large laparotomy is avoided.  A similar approach to extraperioneal revascularization of other distant sites is plausible.

 

64.04 Submucosal Gastric Tumors: Efficacy of a Combined Endoscopic and Laparoscopic Approach

Posted on December 22, 2014

A. C. Pysher1, R. C. Langan1, S. Ram1, S. Morales1, R. S. Jackson1, R. Jha3, N. Haddad2, F. Al-Kawas2, J. Carroll2, P. G. Jackson1  1MedStar Georgetown University Hospital,Department Of General Surgery,Washington, DC, USA 2MedStar Georgetown University Hospital,Division Of Gastroenterology,Washington, DC, USA 3MedStar Georgetown University Hospital,Department Of Radiology,Washington, DC, USA

Introduction:  Gastric submucosal tumors (SMTs) display a wide spectrum of pathologic and prognostic characteristics ranging from benign to highly malignant. Laparoscopic gastric wedge resections have become a commonly utilized treatment method. However, one of the challenges posed by use of the laparoscopic technique alone, is tumor identification. In this study, we provide a comprehensive analysis of SMTs at our institution and present a novel combined laparoscopic and endoscopic resection technique.

Methods:  A retrospective review of a prospectively maintained SMT database was performed which identified 32 patients with gastric submucosal tumors who underwent a combined laparoscopic and endoscopic resection between 2006 and 2010.  All clinicopatholgic data was analyzed.

Results:  Pre-operative evaluation included upper endoscopy and endoscopic ultrasound with biopsy of visible lesions in all of patients. Operative technique utilized intraoperative endoscopic visualization of the lesion, followed by laparoscopic gastric wedge resection under direct endoscopic visualization. Utilizing the combined endoscopic and laparoscopic technique, SMTs were correctly identified in all cases. Final pathology revealed an R0 resection in 97% of patients (31/32). Mean tumor size was 2.48 cm (range, 0.6 to 6 cm) and final pathology revealed gastrointestinal stromal tumors (GISTs) in 18/32 cases. No intra-operative complications occurred. Post-operative course was uncomplicated in 29/32 patients. 3/32 patients experienced early post-operative complications, which included delayed gastric emptying, ileus, and small bowel obstruction. There were no perioperative deaths. 

Conclusion: In one of the first series reporting a combined endoscopic and laparoscopic technique for SMTs, we found heightened identification of all SMTs not afforded by laparoscopy alone. This approach may have spared patients an open resection. Although we found this technique to be safe and efficacious, further studies should assess this novel approach using large nationally representative databases. 

 

62.05 Ex-vivo Normothermic Perfusion (EVNP) to Assess and Repair Kidney Ischemia-Reperfusion Injury

Posted on December 22, 2014

I. Palma1, J. Woloszyn1, R. Abbott1, Y. Smolin1, R. Ramsamooj2, N. Tran2, C. Santhanakrishnan1, R. V. Perez1  1University Of California – Davis,Surgery,Sacramento, CA, USA 2University Of California – Davis,Pathology And Laboratory Medicine,Sacramento, CA, USA

Introduction: Hypothermic storage of organs has been a gold standard for organ preservation in transplantation. However, recent studies have shown that EVNP has the potential to repair and assess the viability of marginal organs prior to transplantation by restoring normal metabolism. The optimal perfusion solution for these functions has not been determined.

Methods: Paired high risk human kidneys (n=8) deemed unsuitable for transplantation were placed on 3 hours of pressure dependent EVNP with a centrifugal pump at 37°C. Kidney was oxygenated via a hollow fiber membrane oxygenator and supplemented with parenteral nutrition & insulin. Kidneys were perfused with leukocyte depleted packed red blood cells (PRBC) as an optimal “repair” solution or whole blood (WB) to more accurately assess viability during early reperfusion.  Exogenous creatinine (0.06 g) was added to the system to assess glomerular clearance.  Pump parameters and urine output were monitored.  Blood and urine samples were collected at the start and every 30 minutes and analyzed for pH, oxygen, electrolytes, creatinine, lactate and neutrophil gelatinase associated lipocalin (NGAL) using a Triage point of care machine. NGAL is an acute kidney injury biomarker. Results were compared between groups using a general linear model for repeated measures test.

Results:The mean age of donors was 60 years. The mean static cold ischemia time was 52.5 hours. The Maryland Aggregate Pathology Index and Kidney Donor Prognostic Index were equal in both groups (5.25 and 87% respectively). Hemodynamic parameters evaluated were flow and resistance, and neither showed statistical significance (p=0.19 and p=0.20). However, PRBC showed a better trend in both parameters. Functional parameters appeared more favorable in the PRBC group but only achieved statistical significance with urine NGAL as shown in Table 1. 

Conclusion:EVNP of high-risk kidneys is possible and combined with POC testing offer a means of assessing organs in real time, potentially making it possible to transplant organs that previously were thought to be untransplantable.  Perfusion with PRBC may have a beneficial effect on the kidney, but may not accurately assess the degree of ischemia-reperfusion injury when compared to the more physiologic WB perfusion. More studies are warranted to determine the contributing factors and clinical usefulness of urinary biomarkers such as NGAL during EVNP.
 

62.06 Relationship of Calcium Sensing and Angiotensin Receptors in the Small Bowel of Mice.

Posted on December 22, 2014

R. Patron-Lozano1, M. I. Rodriguez-Davalos1,2, A. Munoz-Abraham2, J. P. Geibel1,2  1Yale University School Of Medicine,Surgery,New Haven, CT, USA 2Yale University School Of Medicine,Transplantation/Surgery,New Haven, CT, USA

Introduction: The calcium sensing receptor (CaSR) has been identified and its functions described along the entire gastrointestinal tract. Recently, angiotensin receptors have been identified in the small bowel but their physiological function remains controversial. Since both systems are implicated in the regulation and modulation of transmembrane transport of fluids and electrolytes, and even inflammatory diseases, we chose to investigate if there was a connection between these two receptor classes. 

Materials &

Methods:  We used CaSR-/- and Gcmc2 double knockout mice and heterozygote (HET) mice for control. All mice were fasted for 18 hours to reduce intestinal residue; total body weight ranged between 23 and 50 grams. The mice were anesthetized with isoflurane and euthanized. We took 12cm of distal ileum and stripped it from its mesentery. The intestinal lumen was flushed with lactated ringer’s solution in order to remove any remaining intestinal contents. The intestinal loop was then attached to a custom perfusion chamber where it was filled with 2mL of angiotensin at a concentration of 1×10-11 M and weighed. This measurement was determined as weight at t0. The chamber was then filled with lactated ringer’s at 37C and connected to a roller pump which continuously exchanged the fluid and maintained a constant temperature. The experimental time period was 1 hour. The chamber was then emptied and weighed again; this measurement being t1. 

Results: For the knockout mice we observed a significant increase in weight after the administration of angiotensin at 10-11 M concentration between t0 and t1. This difference was consistent in every experiment. The mean weight difference was 73mg or a 3.14% increment in graft weight. For the control HET samples, the weight diminished consistently at t1 96mg or 3.31% of the initial tissue weight.

Conclusion: After performing these experiments, we conclude that there is a functional relationship between angiotensin receptors and calcium sensing receptors in the small bowel of mice. By recording a difference in graft weight after the administration of angiotensin at a concentration of 10-11 M in CaSR -/-;Gcmc2-/- mice we observed that although the angiotensin receptors might be present and stimulated, the absence of CaSR prevents normal physiologic function. Furthermore, by conducting a control experiment with the HET mice, we demonstrated that functional calcium sensing receptors aid the normal physiology of fluid exchange in the small intestine in the presence of angiotensin. These findings can further help us comprehend the extent of the fluid and electrolyte regulators in the body thus providing novel targets for control of hypertension and other diseases.
 

62.07 Innate lymphoid cells in critical illness: is interleukin-33 (IL-33) a potential marker of sepsis?

Posted on December 22, 2014

T. T. Chun1, D. S. Heffernan1, N. Hutchins1, W. G. Cioffi1, C. Chung1, A. Ayala1  1Brown University School Of Medicine,Surgery,Providence, RI, USA

Introduction:  Sepsis remains a major clinical challenge with few effective therapeutic options. The recently described innate lymphoid cells (ILCs) play an important role in sepsis. Specifically, IL-33 mediated group 2 ILCs (ILC2s) produce Th-2 associated cytokines and have a protective effect against sepsis. IL-33 is also known to interact with invariant natural kill T (iNKT) cells to release protective cytokines. Previous studies have demonstrated that ILC2s are implicated in lung inflammation, and sepsis leads to an increased IL-33 level in lung tissue homogenates. We hypothesized that IL-33 is similarly released during sepsis into the blood, acting on the gastrointestinal tract and the liver to stimulate ILC2s as well as iNKT cells. The purpose of this study was to evaluate IL-33 as a marker of sepsis in an experimental model and human subjects.

Methods:  Cecal ligation and puncture (CLP) was performed in both wildtype (WT) and invariant natural killer T cell knockout (iNKT-/-) mice. IL-33 levels were measured in liver tissue homogenates, the serum and the peritoneal fluid using enzyme-linked immunosorbent assays (ELISA). Liver non-parenchymal cells were isolated and stained with antibodies to determine expression of IL-33 receptor (IL-33R). We also obtained blood from septic patients and compared their serum IL-33 levels with those collected from otherwise healthy volunteers.

Results: CLP induced elevation of liver IL-33 in both WT and iNKT-/- mice (p<0.05). Furthermore, following CLP, there was a significantly increased percentage of IL-33R positive cells in WT (2.8 vs 20.9%;p=0.038). Sepsis did not induce an increase in IL-33R expression in NKT-/- mice. There was a trend toward increase in serum IL-33 levels in WT CLP versus sham (43.3 vs 104.6pg/ml;p=0.15), a finding not observed in iNKT-/- mice. Specifically, iNKT-/- mice compared to WT demonstrated significantly lower serum IL-33 levels following CLP (3.6 vs 104pg/ml;p=0.006). Within the peritoneal cavity, CLP induced significantly higher IL-33 levels in WT mice (0.90 vs 10.8pg/ml;p=0.039). In iNKT-/- mice, this alteration of peritoneal IL-33 levels was magnified (3.6 vs 23.4pg/ml;p=0.49). In humans, patients with sepsis had significantly higher serum IL-33 levels than the normal control group (0.17 vs 1.12pg/ml;p=0.0021).

Conclusion: Together, the observations that mouse liver, serum and peritoneal IL-33 levels increase with CLP, as does non-parenchymal cell IL-33R expression, imply that the ILC2s are involved and activated by sepsis. Moreover, this may be effected by the interaction with iNKT cells. Finally, that such changes are not restricted to mice but are also evident in humans, points at novel therapeutic/pathological targets.

 

62.08 Warm Ischemia-Reperfusion Injury Causes Epithelial to Mesenchymal Transformation in Human Cholangiocytes

Posted on December 22, 2014

J. Zhang1, N. Singh1, W. A. Dorsett-Martin1, C. D. Anderson1, T. M. Earl1  1University Of Mississippi,Division Of Transplant And Hepatobiliary Surgery, Department Of Surgery,Jackson, MS, USA

Introduction: The discrepancy between need and supply of liver allografts has led to alternative strategies to increase the organ pool.  One strategy, utilization of donation after cardiac death (DCD) donors, is severely limited by intolerance of the biliary system to warm ischemia resulting in ischemic cholangiopathy which reduces longevity of these allografts.  We hypothesize that warm ischemia followed by cold storage and subsequent reperfusion injury results in epithelial to mesenchymal transformation (EMT) and increased extra-cellular matrix production in human cholangiocytes.

Methods: Primary human cholangiocytes were exposed to no treatment (NT), hypoxic storage at 4°C in Histidine-tryptophan-ketoglutarate (HTK) solution for 7 hours (CIR group) or 3 hours hypoxic normothermic storage in PBS followed by 4 hours hypoxic storage in HTK at 4°C (WIR group).  Following 7 hours hypoxic storage (CIR and WIR), cholangiocytes were reperfused with media under normoxic, normothermic conditions.  Cells were harvested at 24, 72 and 144 hours post-reperfusion.  Quantitative real-time PCR for cholangiocyte epithelial marker AQP1, mesenchymal cell markers αSMA and S100α4, and COLA1 was performed at each time point and normalized to house-keeping gene 18S.

Results: One hundred forty-four hours after reperfusion, AQP1 expression is significantly diminished for WIR (WIR 6.2±0.3 vs. NT 210.6±33.8; p<0.001) but not after CIR (CIR 182.4±28.7 vs. NT 210.6±33.8 vs.; p=0.33) and was significantly lower in WIR compared to CIR (WIR 6.2±0.3 vs. CIR 182.4±28.7; p<0.001).  Interestingly, following CIR cholangiocytes lost Aqp1 mRNA at 24 and 72 hours but regained it by 144 hours.  Mesenchymal marker αSMA mRNA was significantly increased at 144 hours post-reperfusion in cholangiocytes exposed to WIR (NT 2.80±.3 vs. WIR 47.2±2.7; p<0.001), but decreased following CIR (NT 2.8±.3  vs. CIR 1.5±0.6;  p=0.02). This difference was not seen at 24 and 72 hours.  COL1A1 mRNA increased dramatically at 144 hours following WIR but not after CIR (WIR 130.0±6.0 vs. CIR 5.1±0.3; p<0.001), expression following CIR injury did not differ from NT group at any timepoint.  S100α4 trended toward significantly higher expression 144 hours following WIR (WIR 6.0±1.6, CIR 2.8±1.3; p=0.059).

Conclusion: Following warm-ischemia with subsequent cold storage and reperfusion injury human cholangiocytes lose epithelial cell markers, gain mesenchymal cell markers, and have increase expression of collagen 1 mRNA.  Further insight into this process may result in better utilization and success of DCD liver allografts. 

 

62.09 Angiotensin Receptors In The Small Bowel Of Rats: A Novel Target For Hypertension

Posted on December 22, 2014

R. Patron-Lozano2, M. I. Rodriguez-Davalos1, A. Munoz-Abraham1, J. P. Geibel1,2  1Yale University School Of Medicine,Transplantation/Surgery,New Haven, CT, USA 2Yale University School Of Medicine,Surgery,New Haven, CT, USA

Introduction:  The renin-angiotensin-aldosterone system, through angiotensin II (AII), is one of the most important blood pressure regulators. The vasoconstriction caused by AII and the sodium balance effected by aldosterone have been a key target for drugs aiming to control hypertension. Angiotensin (AT) receptors have been well identified in vascular smooth muscle and renal tissue. Moreover, they have also been identified throughout the enteric epithelium, although mainly as a mitogenic agent. In the colon, the localization of functional angiotensin receptors has recently been described by our group and shows a promising potential future target for antihypertensive agents. In this study, we aimed to identify the presence of functional angiotensin receptors using intact small bowel of rats to determine if AII and AT1 receptor agents can modulate small bowel salt transport.

Materials and

Methods:  We used Sprague Dawley rats (280 and 360 grams) fasted for 18 hours, in order to reduce intestinal residue. The rats were anesthetized with isoflurane and euthanized. We took 12cm of distal ileum and stripped it from its mesentery. The intestinal lumen was flushed with lactated ringer’s solution in order to remove any remaining intestinal contents. The intestinal loop was then attached to a custom perfusion chamber where it was filled with 2mL of angiotensin at a concentration of 1×10-11 M and weighed. This measurement was determined as weight at t0. The chamber was then filled with lactated ringer’s at 37C and connected to a roller pump which continuously exchanged the fluid and maintained constant temperature for 1 hour. Then, the chamber was emptied and weighed again; this measurement being t1. For control purposes, the experiment was run using the same technique but the intestinal lumen filled with 2mL of lactated ringer’s. 

Results: We observed a significant decrease in weight after the administration of angiotensin at 10-11 M concentration between t0 and t1. This difference was consistent in every experiment. The mean weight difference was 93mg or 3.20% reduction in net graft weight. For the control samples, the weight remained the same at t1 or even slightly increased by almost 2% compared to the initial tissue weight.

Conclusion: We conclude that there are functional angiotensin receptors in rat small bowel that are sensitive to low dose AII(10-11M). By recording a difference in graft weight after the administration of low dose AII we proved that these receptors are not only present, but they can be stimulated provided adequate characteristics. These findings can further help us comprehend the extent of the fluid and electrolyte regulators in the body thus providing novel targets for control of hypertension and other diseases. 

 

62.10 Donor Plasma Effects On Platelet Function

Posted on December 22, 2014

A. G. Grand1, J. C. Cardenas1, L. Baer1, N. Matijevic1, B. A. Cotton1, J. B. Holcomb1, C. E. Wade1  1University Of Texas Health Science Center At Houston,Department Of Surgery,Houston, TX, USA

Introduction: We and others have demonstrated a significant decrease in platelet count and function within 2 to 3 hours following admission in severely injured trauma patients after transfusion. The decrease in platelet function is in part due to the reduction in platelet count. However, the decrease in function does not always equal the decrease in count observed in trauma patients. We investigated whether transfusion of plasma was another factor contributing to platelet hypofunction following trauma.

Methods: Whole blood samples were taken from healthy volunteers and their baseline platelet function assessed by impedance aggregometry in response to ADP, collagen, thrombin receptor-activating peptide (TRAP), arachidonic acid (AA) and ristocetin using Multiplate Analyzer. Blood samples were then diluted by 30% using the volunteer’s autologous plasma, autologous plasma that was snap frozen and thawed (autologous FFP), and donor fresh frozen plasma (FFP) from Gulf Coast Regional Blood Center. The percent change in platelet function compared to whole blood or autologous FFP was calculated. FFP from five different donors were used to obtain the average change in function. A student’s t test with significance set at p<0.05 was used to determine if dilution, freezing and the use of donor FFP had an effect on platelet function.

Results: Dilution of whole blood with autologous plasma by 30% showed a significant decrease of 30% in platelet function in response to all agonists, as expected, with the exception of TRAP. Autologous FFP had no additional effect on platelet function, with the exception of a reduction in TRAP (p=0.007). Single donor plasma demonstrated a further reduction in ADP (p=0.02), collagen (p=0.006) and AA (p=0.007) compared to autologous FFP. Finally, comparison across multiple donors (n=5) demonstrated a trending, although not significant, reduction in platelet function in response to all agonists with the exception of ristocetin, which remained unchanged.

Conclusion: Dilution is the major contributor in the decrease in platelet function however, donor plasma may have additional negative effects on platelet function following transfusion.  

 

62.11 Current and Future Approaches to the Proteomic Analysis of Traumatic Coagulopathy

Posted on December 22, 2014

C. C. McCoy1, E. Benrashid1, M. L. Shapiro1, S. N. Vaslef1, J. H. Lawson1  1Duke University Medical Center,Department Of Surgery,Durham, NC, USA

Introduction:
Current translational research in traumatic coagulopathy is limited by the heterogeneity of individual injuries and the acute, unpredictable nature of trauma incidence.  As a result, coagulation studies of plasma from trauma patients lack baseline comparisons and are biased by numerous clinical factors including multisystem trauma, pre-hospital resuscitation and blood component administration.  Developing both animal and human models of trauma will permit a more rigorous characterization of coagulation changes resulting from specific organ injury.

Methods:
A mouse model of blunt traumatic brain injury (TBI) was utilized to create a pilot study of TBI-specific coagulation changes.  Three mice received piston-based, blunt TBI (6.8 m/s, 3mm deflection) following anesthesia and scalp retraction, while three mice received anesthesia and scalp retraction alone.  Thirty minutes following intervention, blood was obtained and a proteomic analysis of coagulation was performed.  2-dimensional difference gel electrophoresis (DIGE) was coupled with matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry to identify proteins with altered concentration in an unbiased fashion.

Results:
Multiple coagulation-related proteins were detected by DIGE, and three known coagulation proteins demonstrated concentration alterations (greater than 5% concentration change) following trauma.  Five novel proteins of unknown function also demonstrated alterations in concentration.  Although previously uncharacterized, these proteins could serve as targets for future investigation of injury-induced changes in the plasma proteome.

Conclusion:
Detecting concentration changes of known coagulation factors and novel proteins during TBI reinforces the value of organ-specific models of trauma as a mechanism to study the plasma proteome during injury.  To validate animal model findings, proteomic analyses will be performed on human plasma collected before, during and after elective surgery to develop human, organ-specific injury models.  Data from such research will enhance our knowledge of coagulation changes during tissue trauma, the influence of inflammatory pathways, and their relationship to organ-specific injury.

62.12 Choice of Induction Immunosuppression Influences Net Benefit of Transplant in Pancreas Subcategories

Posted on December 22, 2014

D. J. Taraskiewicz1, D. J. Taber1, S. Nadig1, J. McGillicuddy1, K. D. Chavin1, P. K. Baliga1, C. F. Bratton1  1Medical University Of South Carolina,Charleston, Sc, USA

Introduction: Pancreas transplantation (PTX) is the only treatment which reestablishes the euglycemic state in patients with diabetes.  Success of PTX is predicated upon minimizing deleterious infection and rejection episodes.  Induction immunosuppression (II) including IL2 receptor antagonists (IL2) and depleting antibodies (DA) are utilized to attenuate rejection in PTX in 90% of cases nationally; however, data regarding optimal II modality in PTX is inadequate. The aim of this analysis was to examine the risks and benefits of II compared across PTX type:  simultaneous PTX (SPK) vs. Pancreas after kidney (PAK) or pancreas transplant alone (PTA).

Methods: This was a respective analysis of PTX performed at MUSC between 2000-2014 using medical records to determine one year rates of infection, rejection, graft loss, and patient death for IL2 vs DA II, with sub-analyzes by ethnicity.   Data included baseline donor and patient characteristics, univariate factors were evaluated with SPSS v22. Factors demonstrating significant differences were included in multivariate modeling.  Patients with graft loss or mortality within the first month due to surgical complications were excluded.

Results: 205 PTX were included (154 [75%] SPK, 51 [25%] PAK/PTA); 1 yr infection rates were significantly higher for SPKs induced with DA vs. IL2 (41% vs 25%; p=0.033), while1 yr infection rates for PAK/PTA trended towards significance (41% DA vs. 21% IL2; p=0.202).    There was no difference in rejection rates in SPKs (10% DA vs. 16% IL2; p=0.355).  However, in the PAK/ PTA group, DA use demonstrated a trends towards improvement in 1 yr rejection (27% DA vs. 43% IL2; p=0.277), and graft loss (5.4% DA vs. 21.4% IL2; p=0.086).  II in African Americans (AA) receiving PTX did not influence infections (46% vs. 36%; p=0.357) or rejections (15% DA vs. 13% IL2; p=0.745).

Conclusion: These results suggest that the efficacious use of II in PTX is dependent on transplant type, with SPKs benefiting more from IL2, as evident by lower infection rates without differences in the rejection risk.  In contrast, PAK/PTAs appear to benefit from DA.  Although ethnicity may influence outcomes in PTX, II efficacy was not dramatically different based on recipient ethnicity.

 

62.13 Normothermic Extracorporeal Liver Perfusion for Extended Criteria Livers

Posted on December 22, 2014

B. Banan1, H. Chung1, Z. Xiao1, Y. Tarabishy1, P. Manning2, D. McGraw2, G. Upadhya1, T. Mohanakumar1, Y. Lin1, W. C. Chapman1  1Washington University School Of Medicine In St. Louis,Trasnplant/Surgery/Medicine,St. Louis, MO, USA 2Vasculox, Inc,St. Louis, MO, USA

Introduction: A critical shortage of donor livers has led to increased efforts to expand the criteria for suitable donors.  However, the higher sensitivity of extended criteria livers to ischemia reperfusion injury (IRI) is a major obstacle in organ cold preservation which jeopardizes post-transplant graft viability. Normothermic extracorporeal liver perfusion (NELP) has been investigated to reduce ischemic damage, restore physiologic function and enable surgeons to assess viability of the liver prior to transplant. The goal of this study was to investigate the efficacy of NELP to re-establish physiologic parameters and metabolic functions in livers procured after an extended warm ischemia time (WIT) in a porcine model. We for the first time compared physiological parameters of livers maintained continuously on NELP to ones which preserved in cold solution.

Methods: Livers from 9 female landrace pigs (30-34kg) were subjected to 20 min (WIT-20 group, n=3), 40 min (WIT-40 group, n=3) and 60 min (WIT-60 group, n=3) warm ischemia followed by 8 hours of NELP.  To assess superiority of NELP over cold storage, WIT-40 group was compared to three livers subjected to 40 min warm ischemia time followed by 6 hours of cold storage (WCIT-40 group) in HTK solution and 2 hours of NELP. Blood samples and biopsies were taken at hourly intervals. We considered the final 2-hours of NELP as the test phase in all groups (ie, functional reperfusion period) for comparison purposes.  Groups were compared using 2-way ANOVA test, with p<0.05 as significant.

Results: NELP stabilized transaminases and total bilirubin levels in all WIT groups. This was accompanied by significant improvement in bile and albumin production, and a remarkable decline in lactate and INR values. pH, Na+, K+, cl-, pCO2, bicarbonate and glucose measures were stabilized at physiologic levels.  Oxygen extraction ratio was recovered in all WIT groups during first 30 minutes of machine perfusion and maintained until the end of study. Pathology review showed significant improvement from 0 hour (mild to moderate sinusoidal dilation and zone 3 necrosis) to end time of the NELP run (minimal hepatocyte necrosis and mild IRI) in all WIT study groups. WIT-40 and WCIT-40 comparison revealed significantly higher values of bile production, albumin and oxygen extraction ratio in WIT-40. However, WCIT-40 group showed higher AST, ALT, lactate and INR levels accompanied by moderate IRI. The following table summarizes some significant results obtained at the end of NELP run.

Conclusion: NELP improves and maintains metabolic and functional parameters of livers with either short or extended warm ischemia times and reduces IRI when compared to livers subjected to cold ischemia time.

 

62.14 Complete Thymectomy in Adult Rats with Non-invasive Endotracheal Intubation

Posted on December 22, 2014

V. R. Rendell3, J. Lin2, M. L. Markert2, T. V. Brennan1  1Duke University Medical Center,Surgery,Durham, NC, USA 2Duke University Medical Center,Pediatrics,Durham, NC, USA 3Duke University Medical Center,Medicine,Durham, NC, USA

Introduction:  Rodent thymectomy enabled defining the role of the thymus in lymphocyte differentiation, self-tolerance, and immunotolerance in the setting of allograft transplantation and tumor metastasis.  The technique facilitates studies involving T-cell depletion or the adoptive transfer of defined T cell populations without the re-emergence of native naïve T cells. Thymectomy in adult rats has proven to be more challenging than neonatal rat thymectomy due to increased rates of bleeding, tracheal injury and pneumothorax.  The objective of the present study was to develop a technique for adult rat thymectomy with low complication rates and reliable complete removal of the thymus.

Methods:   The protocol was evaluated for 26 rats. An endotracheal intubation cannula was fashioned from an angiocatheter needle. Rat intubation was accomplished using trans-cutaneous tracheal illumination to visualize the aperture of the larynx prior to insertion of the prepared cannula within the angiocatheter tubing. A minimally invasive 2 cm skin incision was followed by a 1.5 cm median sternotomy and division of the pre-tracheal muscles. The inferior thyroid lobes were carefully dissected away from the pleural lining. After complete removal of the thymus, a three layer closure with surgical glue application was performed.  Removed tissue was stained by H&E and with cytokeratin staining. Peripheral blood from the rats was analyzed for persistent depletion of naïve T cells post-operatively by staining with naïve T cell markers and dividing the percent of each cell type present by the absolute lymphocyte count. 

Results: The mean intubation time was 45sec +/- 5 sec, and her mean operation time was 15 ± 3 min. There was an associated 96% immediate post-operative survival rate.  There was no intraoperative mortality. Twenty-four rats recovered fully. One rat survived after re-exploration of the chest.  All removed thymus tissue was submitted to path. Immunohistochemistry cytokeratin would help differentiate thymic tissue from lymph nodes. All thymectomies performed were confirmed to be complete. Rats who underwent thymectomy had minimal new naïve type T cells post-operatively. The thymectomized rats maintain overall T cell counts but have diminished naïve T cells. 

Conclusion: This method is appropriate when complete thymectomy with minimal complications is desired for further immunological studies in athymic adult rats. All thymectomies were complete as evidenced by immunohistochemical (IHC) staining of mediastinal tissue, and absence of naïve T-cells by flow cytometry, and the procedure had a 96% survival rate.

 

62.16 Small Intestine Microbiota Is Altered In Short Bowel Syndrome in the Zebrafish.

Posted on December 22, 2014

K. A. Schall1, J. W. Debelius3, K. A. Holoyda1, R. Knight3, T. C. Grikscheit1,2  2Keck School Of Medicine,Los Angeles, CA, USA 3University Of Colorado Boulder,Department Of Chemistry And Biochemistry,Boulder, CO, USA 1Children’s Hospital Los Angeles,Department Of Pediatric Surgery,Los Angeles, CA, USA

Introduction:
Short bowel syndrome (SBS) is characterized by malnutrition, mucosal inflammation and small intestine bacterial overgrowth (SIBO). The definition and evaluation of SIBO is poorly understood but hypothesized to arise from stasis of luminal contents and colonization from the colon.  In some cases, prophylactic antibiotics are given. Previous studies characterized only fecal and colonic mucosal samples, but the small intestinal milieu is known to be quite different. We recently developed a novel model of SBS in the zebrafish: a high proximal stoma is formed at the fish correlate to the human jejunum, with a distal Hartman’s pouch.  As in the human disease, weight loss and intestinal dilation are noted.  We aimed to characterize the small intestine microbiome alterations associated with SBS in our novel zebrafish SBS model.

Methods:
The SBS surgery consisted of a ventral laparotomy, proximal stoma creation, distal bowel ligation and removal of middle portion of intestine to prevent fistula formation. Nineteen adult male zebrafish underwent SBS surgery with proximal ostomy and distal ligation, sham with ventral laparotomy only (n=9), or control with no operation (n=10); half housed separately and together to control for environment. The mid-portion of the intestine was removed at the time of SBS operation while the proximal and distal intestine were harvested from the remaining fish after 2 weeks. DNA was extracted with the PowerSoil kit and 16s rRNA based gene sequencing completed to identify the bacterial species present. Alpha rarefaction to 2500 segments/sample was used to balance loss of samples.  Unweighted Unifrac distance was compared using permanova and anosim test.  Alpha diversity was compared by kruskal wallis.  Paired samples within an individual zebrafish, which consisted of the proximal (“fed”) and distal (“unfed”) intestine, were compared using kruskal wallis.  Qiime was used to generate taxonomy, which was plotted using Excel.

Results:
Surgery increased the operational taxonomic units (OTUs) from the genus Shewanella and decreased OTUs from the family Aeormonadacaea and genus Clocibacterium after SBS surgery as compared to the controls. Environment consisting of the tank water was significantly different than the intestinal bacteria (p=.001), having no effect on intestinal microbiota changes. The paired controls of proximal and distal intestine in SBS, despite relative changes in some OTUs, revealed a core microbiota that is unchanged (p<0.01).

Conclusion:
The microbiota is altered in the small intestine after SBS in a zebrafish model, possibly correlating to the human disease process, leading to increased inflammation and SIBO.  Understanding the luminal factors after surgery, including the microbiome, may indicate novel therapeutic interventions for children with this morbid disease.
 

62.19 TRPA1 Mediates the Effects of Hypothermia on the Monocyte Inflammatory Response

Posted on December 22, 2014

N. J. Galbraith1, A. T. Billeter1, C. Lawson1, J. D. Rice1, H. C. Polk1  1University Of Louisville,Hiram C. Polk Jr MD Department Of Surgery,Louisville, KY, USA

Introduction:

Unintentional hypothermia is a known risk factor for peri-operative septic and cardiovascular complications.  We have previously shown that hypothermia increases the pro-inflammatory response, while the anti-inflammatory response is suppressed via Interleukin 10 (IL-10) and microRNA-155 (miR -155). However, the exact molecular mechanisms by which hypothermia affects the monocyte immune response are unknown. Transient receptor potential ion channels (TRP) are a large group of cell membrane bound receptors, implicated in pain, cancer and sepsis. These channels are activated by a variety of chemical substances or physical stress such as mechanical forces or temperature. Three TRP-channels (TRPA1, TRPM8, and TRPV1) are temperature sensitive of which TRPA1 and TRPM8 sense cold whereas TRPV1 is heat activated. The purpose of this study was to investigate whether these channels mediate the effects of hypothermia on the monocyte inflammatory response.

Methods:
Primary human monocytes were freshly isolated from healthy donors, treated for 24h then stimulated with LPS (100ng/ml) at 32°C or 37°C. Cell culture supernatant and total RNA were isolated. Expression of TRPA1, TRPM8, and TRPV1 mRNA as well as miR-155 was determined using qRT-PCR. Cytokines in the supernatant were measured using ELISAs. Specific inhibitors of TRPA1 (HC- 030031) and a specific activator of TRPV1 (capsaicin) were used to block or activate the specific channels. Statistical analysis was performed using the Wilcoxon signed-rank test. 

Results:
TRPM8 mRNA was not expressed in monocytes whereas TRPA1 and TRPV1 were highly expressed. TRPV1 mRNA-expression was significantly suppressed at 32°C but not at 37°C; TRPA1 was strongly induced at 32°C and 37°C. Immunofluorescence microscopy confirmed that monocytes express TRPA1 and TRPV1 on their surface. Blocking TRPA1 at 32°C tended to decrease TNF secretion after 24h. Similarly, TRPV1 activation at 32°C tended to suppress TNF secretion. By comparison, TRPA1 blockade (93.7 ± 29.8pg/ml vs. 43.6 ± 9pg/ml, p<0.05) and TRPV1 (5.0 ± 6.1pg/ml vs. 43.6 ± 9pg/m, p<0.05) activation at 32°C significantly increased IL-10. In addition, TRPA1 blockade and TRPV1 activation at 32°C both suppressed miR-155 expression p<0.05). 

Conclusion:
These findings suggest that hypothermia mediates its effects on the monocyte inflammatory response through TRP channels. Both the blockade of the cold activated TRPA1 channels, and the activation of the heat activated TRPV1 channels can restore normal cytokine production, thus alleviating the detrimental effects of hypothermia. This modulation of the monocyte inflammatory response may provide critical new treatment options in patients suffering hypothermic associated complications.
 

62.20 Characterizing Myeloid-derived Suppressor Cells by Expression of LIGHT and its Cognate Receptors

Posted on December 22, 2014

J. F. Calata1, S. Jayaraman1, B. S. Prabhakar1, A. V. Maker1  1University Of Illinois At Chicago,Chicago, IL, USA

Introduction:

Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells that negatively regulate immune responses during tumor progression. There remains a significant gap in our understanding of their phenotypical and functional heterogeneity. We have previously demonstrated that the immunostimulatory cytokine LIGHT (TNFSF14) can mature certain myeloid cells, and that its presence in the tumor microenvironment can stimulate an anti-tumor T-cell response.  We, therefore, determined to define the expression of LIGHT and its cognate receptors on MDSCs.

Methods:

Female BALB/c mice, 6-8 weeks of age, were injected subcutaneously with 1×106 CT26 cells (murine colorectal carcinoma cell line).  When tumors reached 1 cubic centimeter, splenectomy was performed.  Splenocytes were isolated and evaluated with flow cytometry for expression CD11b, Ly6G, Ly6C, HVEM, LTBR, and LIGHT.

Results:

Monocytic-MDSC (M-MDSC, CD11b+/Ly6G+) and granulocytic-MDSCs (G-MDSC, CD11b+/Ly6C+) were identified in tumor bearing mice.  LIGHT was expressed preferentially on G-MDSCs (94% vs. 39%, p= 0.0006).  The LIGHT receptors HVEM and LTBR were also expressed on MDSC subsets. LTBR expression was high in both populations, but increased in G-MDSC compared to M-MDSC (p=0.018), while HVEM expression was lower in G-MDSC (84% vs. 40%, p=<0.0001).  Populations of M-MDSC were, therefore, characterized as LIGHTlowLTBRlowHVEMhi while G-MDSC were LIGHThiLTBRhiHVEMlow.

Conclusion:

MDSCs can be characterized by their expression of LIGHT and its cognate receptors.  Based on this data, the effect of LIGHT and LIGHT receptor blockade on MDSC function warrants further investigation since inhibition of immunosuppressive MDSC may be a promising strategy for tumor immunotherapy.

 

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