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Hematopoietic stem cell transplantation from anti-cytomegalovirus immunoglobulin G (anti-CMV-IgG) positive donors facilitated immunological recovery post-transplant, which may indicate that chronic CMV infection has an effect on the immune system. This can be seen in the recipients after reconstitution with donor lymphocytes. We evaluated the composition of lymphocytes at hematologic recovery in 99 patients with hematologic malignancies post hematopoietic stem cell transplantation (HSCT). Anti-CMV-IgG seropositivity of the donor was associated with higher proportions of CD4+ (227.963 ± 304.858 × 106 vs. 102.050 ± 17.247 × 106 cells/L, p = 0.009) and CD4+CD25high (3.456 ± 0.436 × 106 vs. 1.589 ± 0.218 × 106 cells/L, p = 0.003) lymphocytes in the blood at hematologic recovery. The latter parameter exerted a diverse influence on the risk of acute graft-versus-host disease (GvHD) if low (1.483 ± 0.360 × 106 vs. 3.778 ± 0.484 × 106 cells/L, p &lt, 0.001) and de novo chronic GvHD (cGvHD) if high (3.778 ± 0.780 × 106 vs. 2.042 ± 0.261 × 106 cells/L, p = 0.041). Higher values of CD4+ lymphocytes in patients who received transplants from anti-CMV-IgG-positive donors translated into a reduced demand for IgG support (23/63 vs. 19/33, p = 0.048), and these patients also exhibited reduced susceptibility to cytomegalovirus (CMV), Epstein–Barr virus (EBV) and/or human herpes 6 virus (HHV6) infection/reactivation (12/50 vs. 21/47, p = 0.032). Finally, high levels (³0.4%) of CD4+CD25high lymphocytes were significantly associated with better post-transplant survival (56% vs. 38%, four-year survival, p = 0.040). Donors who experience CMV infection/reactivation provide the recipients with lymphocytes, which readily reinforce the recovery of the transplanted patients’ immune system.

Several different surgical repair procedures for symptomatic rectocele have been described with variable results. In our clinic, a modified anterolateral rectopexy is used. In this article we evaluate our results, with emphasis on patient satisfaction. From 2001 until 2003, twenty patients with a symptomatic rectocele were treated by anterolateral rectopexy. The preoperative dynamic defecogram and anorectal complaints were analyzed and compared to postoperative outcome via a standardized questionnaire. After surgery, all rectoceles were restored as shown by postoperative defecogram. Anorectal symptoms (incomplete evacuation, continuous urge, prolapse, digital evacuation) were improved in 40%. As new-onset symptoms, dyspareunia (50%), digital support (55%) and incomplete evacuation (75%) were mentioned frequently. Most of the patients with larger rectoceles (>3.5 cm) had increased anorectal complaints after surgery. Anterolateral rectopexy for treatment of rectocele give limited improvement of anorectal complaints. Besides, many patients developed new complaints postoperatively and hence overall satisfaction was low.

The development of indirect inguinal hernias in infants is caused by a patent processus vaginalis (PPV). Consequently, this type of hernia is cured by simple herniotomy. In adults, however, herniotomy alone is accompanied by a high recurrence rate. This indicates that additional factors play a part in the development of indirect inguinal hernias in adults. The aim of this study was to determine the etiology of the development of an indirect hernia in adult life. Also, the prevalence of a PPV without clinical evidence of a hernia was determined and related to age. From November 1998 until February 2002, 599 patients from four different teaching hospitals, who underwent abdominal laparoscopy for various pathologies, were included. During laparoscopy, the deep inguinal ring was bilaterally inspected. Patients undergoing laparoscopy for inguinal hernia repair were excluded. Mean age was 45 years (range 8–89 years). Thirty-two percent (189/599) were male. Twelve percent (71/599) had PPV, all without clinical symptoms. Fifty-five percent (39/71) with PPV were male (P

Some cancers treated with allogeneic hematopoietic stem cell transplantation (HSCT) are sensitive to natural killer cell (NK) reactivity. NK function depends on activating and inhibitory receptors and is modified by NK education/licensing effect and mediated by coexpression of inhibitory killer-cell immunoglobulin-like receptor (KIR) and its corresponding HLA I ligand. We assessed activating KIR (aKIR)-based HLA I–dependent education capacity in donor NKs in 285 patients with hematological malignancies after HSCT from unrelated donors. We found significantly adverse progression-free survival (PFS) and time to progression (TTP) in patients who received transplant from donors with NKs educated by C1:KIR2DS2/3, C2:KIR2DS1, or Bw4:KIR3DS1 pairs (for PFS: hazard ratio [HR], 1.70; P = .0020, Pcorr = .0039; HR, 1.54; P = .020, Pcorr = .039; HR, 1.51; P = .020, Pcorr = .040; and for TTP: HR, 1.82; P = .049, Pcorr = .096; HR, 1.72; P = .096, Pcorr = .18; and HR, 1.65; P = .11, Pcorr = .20, respectively). Reduced PFS and TTP were significantly dependent on the number of aKIR-based education systems in donors (HR, 1.36; P = .00031, Pcorr = .00062; and HR, 1.43; P = .019, Pcorr = .038). Furthermore, the PFS and TTP were strongly adverse in patients with missing HLA ligand cognate with educating aKIR-HLA pair in donor (HR, 3.25; P = .00022, Pcorr = .00045; and HR, 3.82; P = .027, Pcorr = .054). Together, these data suggest important qualitative and quantitative role of donor NK education via aKIR-cognate HLA ligand pairs in the outcome of HSCT. Avoiding the selection of transplant donors with high numbers of aKIR-HLA-based education systems, especially for recipients with missing cognate ligand, is advisable.

Among cancers treated with allogeneic hematopoietic stem-cell transplantation (HSCT), some are sensitive to natural killer (NK) cell reactivity, described as the "missing self" recognition effect. However, this model disregarded the NK cell licensing effect, which highly increases the NK cell reactivity against tumor and is dependent on the coexpression of inhibitory killer cell immunoglobulin-like receptor (iKIR) and its corresponding HLA Class I ligand. We assessed clinical data, HLA and donor iKIR genotyping in 283 patients with myelo- and lymphoproliferative malignancies who underwent HSCT from unrelated donors. We found dramatically reduced overall survival (OS), progression free survival (PFS), and time to progression (TTP) among patients with malignant diseases with the lack of HLA ligand cognate with this iKIR involved in NK cell licensing in corresponding donor (events 83.3% vs. 39.8%, P = 0.0010; 91.6% vs. 47.7%, P = 0.00010; and 30.0% vs. 17.3%, P = 0.013, for OS, PFS, and TTP, respectively). The extremely adverse PFS have withstand the correction when patient group was restricted to HLA mismatched donor-recipient pairs. The incidence of aGvHD was comparable in two groups of patients. In malignant patients after HSCT the missing HLA ligand for iKIR involved in NK cell licensing in corresponding donor ("missing licensing proof") induced extremely adverse survival of the patients due to the progression of malignancy and not to the aGvHD. Avoiding the selection of HSCT donors with the "missing licensing proof" in the malignant patient is strongly advisable.

Interleukin-17A is a hallmark of a subset of CD4+ lymphocytes called T H 17. Allogeneic hematopoietic stem cell transplantation (HSCT) induces an immune response that facilitates graft acceptance, but if clinically apparent as acute graft-versus-host disease (aGvHD), it may adversely affect transplantation outcomes. T H 17 cells are involved in the inflammatory processes associated with several diseases, including inflammatory bowel disease (IBD) as a prototype. In this study we investigated the presence of IL-17–producing cells among peripheral blood mononuclear cells (PBMC) of patients after HSCT. The 48 patients of median age 45 years (range, 1.0–64 years), experienced hematologic malignancies (n = 45) or nonmalignant disorders (n = 3), treated with matched unrelated (n = 24) or sibling (n = 24) transplants. We examined IL-17–producing cells in alloreactive reactions after HSCT. PBMC were stimulated with BD Leukocyte Activation Cocktail (Ionomycin, Brefeldin A, and phorbol myristic acetate (PMA)) in the presence of BD GolgiStop. After stimulation the cells were labeled with anti-CD4 and intracellular anti–IL-17A monoclonal antibodies. IL-17+ cell proportions were analyzed in the CD4+ lymphocyte gate. We observed that patients at the time of hematologic reconstitution had higher proportions of IL-17–producing cells than healthy control subjects (0.73 ± 0.13 vs 0.19 ± 0.06%; P = .019). Fourteen patients displayed the first symptoms of aGvHD at the time of hematologic reconstitution, when they showed lower proportions of IL-17+ cells among CD4+ lymphocytes than their counterparts lacking aGvHD at a similar time after transplantation (0.29 ± 0.09 vs 0.73 ± 0.13%; P = .024). Eight patients developed aGvHD after hematologic reconstitution (median, 34 days). All of these patients displayed lower proportions of IL-17–producing CD4+ cells on the day of aGvHD compared with their initial measurements preceding this complication (0.34 ± 0.14 vs 1.07 ± 0.37%; P = .01).

Objective: A prospective study was undertaken to assess the interobserver variability in CT scan interpretation in patients with suspected acute appendicitis. Methods: 103 adult patients with suspected acute appendicitis underwent unenhanced helical multislice CT scanning of the abdomen. All patients subsequently underwent laparoscopy by a surgeon who was blind to the diagnosis suggested by the CT scan. All CT scans were interpreted by group A, B and C radiologists with different levels of expertise. Results: Acute appendicitis was diagnosed on multislice CT scans in 69%, 74% and 80% by group A, B, and C radiologists, respectively. At laparoscopy, 83% of patients were diagnosed with acute appendicitis. For group A, B and C radiologists, the specificity of CT scanning for the diagnosis of acute appendicitis was 94%, 94% and 100%, respectively, the sensitivity was 81%, 88% and 95%, the positive predictive value was 98.6%, 98.7% and 100%, and the negative predictive value was 50%, 68% and 81%. The difference in the proportion of positive outcomes between observer groups A and C was significant. During laparoscopy, 12 patients were diagnosed with other diseases. These were all correctly diagnosed by group C radiologists; groups A and B radiologists missed the diagnosis of colitis in one patient. All other disorders were diagnosed correctly. Conclusions: The sensitivity of CT interpretations for the diagnosis of acute appendicitis differs considerably between radiologists. This interobserver variability has to be taken into account during implementation of routine CT scanning in patients with suspected acute appendicitis. Only in centres with expert CT radiologists is the implementation of routine CT scanning in patients with suspected acute appendicitis justified.

Background: Incisional hernia repair can be a significant challenge for both surgeon and patient. Despite the growing amount of literature describing various methods of surgical techniques, little has been published regarding the natural course of incisional hernia and the opinions about indications for incisional hernia repair. Methods: A questionnaire was sent to a group of surgeons internationally renowned in incisional hernia surgery and research. Results: Pain and limitations of daily activities were considered the most important indications for repair. Cosmetic complaints were seen as least important. About 23% of patients were asymptomatic. More than 20% did not receive surgical treatment. Conclusions: A large proportion of patients with incisional hernia is not operated. Despite this large group of patients, valid data describing the natural course are absent. A prospective trial monitoring incisional untreated hernias as well as comparing conservative treatment with repair should be performed.

Background Closed laparoscopy, employing a Veress needle and blind insertion of the first trocar, is favoured by most laparoscopic surgeons. The potential danger of this technique is the occurrence of visceral or vascular injury. Establishment of pneumoperitoneum by an open technique using a blunt-tipped trocar may be a safer alternative. Methods and Results Retrospective review of the literature and the authors' experience was used to compare closed and open laparoscopy. Data on closed laparoscopy in 489 335 patients and on opcn laparoscopy in 12 444 patients were culled. Ratcs of visceral and vascular injury were respectively 0.083 and 0.075 per cent after closed laparoscopy, and 0.048 per cent and zero after open laparoscopy. Mortality rates after closed and open laparoscopy were respectively 0.003 per cent and zero. Pearson X 2 analysis demonstrated a statistically significant difference in terms of visceral and vascular injury between closed and open laparoscopy (P = 0002); there was no such difference for mortality rates. Conclusion Open establishment of pneumoperitoneum is advocated in laparoscopic surgery because it is safer than the closed method.

Background Open cholecystectomy (OC) is often preferred over laparoscopic cholecystectomy (LC) in patients with liver cirrhosis and portal hypertension, but evidence is lacking to support this practice. This meta-analysis aimed to clarify which surgical technique is preferable for symptomatic cholecystolithiasis in patients with liver cirrhosis. Methods A meta-analysis was conducted according to the PRISMA guidelines. Articles published between January 1990 and October 2011 were identified from MEDLINE, Embase and the Cochrane Library. Randomized clinical trials (RCTs) comparing outcomes of OC versus LC for cholecystolithiasis in patients with liver cirrhosis were included. The quality of the RCTs was assessed using the Jadad criteria. Results Following review of 1422 papers by title and abstract, a meta-analysis was conducted of four RCTs comprising 234 surgical patients. They provided evidence of at least level 2b on the Oxford Level of Evidence Scale, but scored poorly according to the Jadad criteria. Some 97·0 per cent of the patients had Child–Turcotte–Pugh (CTP) grade A or B liver cirrhosis. In all, 96·6 per cent underwent elective surgery. No postoperative deaths were reported. LC was associated with fewer postoperative complications (risk ratio 0·52, 95 per cent confidence interval (c.i.) 0·29 to 0·92; P = 0·03), a shorter hospital stay (mean difference − 3·05 (95 per cent c.i. − 4·09 to − 2·01) days; P < 0·001) and quicker resumption of a normal diet (mean difference − 27·48 (−30·96 to − 23·99) h; P < 0·001). Conclusion Patients with CTP grade A or B liver cirrhosis who undergo LC for symptomatic cholecystolithiasis have fewer overall postoperative complications, a shorter hospital stay and resume a normal diet more quickly than those who undergo OC.

Background Incarceration of inguinal, umbilical and cicatricial hernias is a frequent problem. However, little is known about the relationship between the use of mesh and outcome after surgery. The goal of this study was to describe the relationship between the use of mesh in incarcerated hernia and the clinical outcome. Patients and methods Correspondence, operation reports and patient files between January 1995 and December 2005 of patients presented at one academic and one teaching hospital in Rotterdam were searched for the following keywords: incarceration, strangulation and hernia. The patient characteristics, clinical presentation, pre-operative findings and clinical course were scored and analysed. Results A total of 203 patients could be identified: 76 inguinal, 52 umbilical, 39 incisional, 14 epigastric, 14 femoral, five trocar and three spigelian hernias. In the statistical analysis, epigastric, femoral, trocar and spigelian hernias were pooled, due to their small group sizes. One patient was excluded from the analysis because the hernia was not corrected during operation. In total, 99 hernias were repaired using mesh versus 103 primary suture repairs. Twenty-five wound infections were registered (12.3%). One mesh was removed during a reintervention for anastomotic leakage, although no signs of wound infection were present. Nine patients died, none of them due to wound-related problems [one cardiovascular, one ruptured aneurysm, two anastomotic leakage, two sepsis e causa incognita (e.c.i.), three pulmonary complications]. Univariate analysis showed that female patients (P = 0.007), adipose patients (P = 0.016), patients with an umbilical hernia (P = 0.01) and patients who underwent a bowel resection (P = 0.015) had a significantly higher rate of wound infections. The type of repair (e.g. primary suture or mesh), use of antibiotic prophylaxis, gender, ASA class and age showed no significant relation with post-operative wound infection. After logistic regression analysis, only bowel resection (P = 0.020) showed a significant relation with post-operative wound infection. Conclusions Wound infection rates are high after the correction of acute hernia, but clinical consequences are relatively low. Mesh correction of an acute hernia seems to be safe and should be considered in every incarcerated hernia.

In order to create a pneumoperitoneum with the Veress needle, it is generally advocated that the abdominal wall should be lifted. Lifting is aimed at increasing the distance between the abdominal wall and the intraabdominal structures. This study was conducted to compare lifting (L) and nonlifting (NL) of the abdominal wall.All patients scheduled for laparoscopic surgery without previous abdominal surgery or morbid obesity were included in the study group. The number of attempts needed for proper positioning of the needle was assessed.A total of 150 patients were randomized. There were no complications. The number of attempts needed for correct positioning of the Veress needle was significantly higher in the L group than in the NL group (31 of 75 vs nine of 75, p0.001). The body mass index (BMI) of patients in whom peritoneal entry needed more than one puncture was significantly higher than the BMI of patients with immediate proper placement (28.3 vs 24.7 kg/m(2), p0.05).Abdominal wall lifting is not necessary.

Background and objectives: A semi-quantitative Real-Time PCR strategy was developed to identify indicator organisms for an indication on anastomotic leakage in peritoneal drainage fluid, Escherichia coli and Enterococcus faecalis. Materials and Methods: The analytical performance of the amplification method was validated with 10 culture-positive and 7 culture negative peritoneal drain fluid samples, obtained from 9 different patients with a colorectal anastomosis. Results: Real-Time PCR results were fully concordant with the microbiological culture results. However, among the culture negative samples, four false-positive RT-PCR results were found. All false-positives originated from a single patient with a surgical site infection. This may indicate an elevated sensitivity of the RT-PCR method. Conclusion: The results showed that the semi-quantitative RT-PCR method has the clear potential to be useful as a powerful tool in early detection of anastomotic leakage.

Background Cholecystectomy is a common operation with a low mortality rate. Although serious complications are rare, it is important to consider them in patients with persisting symptoms after cholecystectomy. We describe a patient with jaundice after laparoscopic cholecystectomy, which was based on a pseudoaneurysm of the cystic artery. Case outline A 68-year-old man was admitted with persisting jaundice three months after laparoscopic cholecystectomy. Colour-Doppler imaging showed pulsatile waves in a 3 cm diameter cystic mass compressing the common bile duct. Selective angiography showed an aneurysm of the cystic artery. An endoluminal prosthesis was placed at ERCP. Results Because of occlusion the stent was changed, but the patient then sustained massive intestinal bleeding. The false aneurysm which had developed from an open remnant of the cystic artery had ruptured into the common bile duct. At laparotomy the cystic artery was succesfully ligated. The patient recovered completely. Discussion There are only four reports of a pseudoaneurysm complicating cholecystectomy, two after open cholecystectomy and two after laparoscopic cholecystectomy. This is the first case reporting a false aneurysm of the cystic artery after cholecystectomy in which the patient presented with jaundice. It shows that this complication can be treated accurately provided that the diagnosis is made early.

In view of constructing a gastric tube after esophagus resection, the vascular anatomy of the greater curvature of the stomach, especially the connection between the left and right gastro-epiploic arteries, was investigated. The vascular anatomy was studied in 20 embalmed human specimens. After dissection a gastric tube of 4 cm wide was constructed, using the greater gastric curvature. Various lengths of the arterial arcades were measured. In 70% an anastomosis between the right and left gastro-epiploic arteries was present. With the construction of an isoperistaltic gastric tube, in which the left gastro-epiploic artery is left in situ (ligating it at the splenic hilus), there is an 18.7% increase of length of arterial arcade along the gastric tube. Leaving the left gastro-epiploic artery in situ increases the feeding arterial arcaded-length along the gastric tube with 5.0 cm (19%).

Background: Pain syndromes of somatic and neuropathic origin are considered to be the main causes of chronic pain after open inguinal hernia repair. Nerve-identification during open hernia repair is suggested to be associated with less postoperative chronic pain. The aim of this study was to define clinically relevant surgical anatomical zones facilitating efficient identification of the three inguinal nerves during open herniorrhaphy. Method: Through dissection of 18 inguinal areas of embalmed and unembalmed human cadavers, identification zones were developed for the inguinal nerves (in particular for the genital branch of the genitofemoral nerve). Results: The iliohypogastric nerve was identifiable running approximately horizontally and ventrally to the internal oblique muscle perforating the external oblique aponeurosis at a mean of 3.8 cm (range 2.5-5.5 cm) cranially from the external ring. When present, the ilioinguinal nerve was identifiable running ventrally and parallel to the spermatic cord, dorsally from the aponeurosis of the external oblique muscle. Identification of the genital branch of the genitofemoral nerve was more comprehensive. The course of the genital branch is laterocaudal at the level of the internal inguinal ring. Conclusion: Based on the newly defined identification zones, peroperative identification of all inguinal nerves is possible. Further research is warranted to assess clinical feasibility of these zones and to evaluate the influence of (facultative) division, preservation or omittance of the identification of inguinal nerves on the incidence of chronic pain.

Normal karyotype AML patients with FLT3 ITD have a higher risk of relapse than those lacking this mutation (Estey EH, Am J Hematol. 2013). Our comparative genomic hybridization study (77 patients) showed that AML cases with as compared to those without FLT3 ITD had significantly lower number of amplifications or deletions (number of total CNV aberrations: 18±4 vs 51±8, p=0.003). The latter observation in concert with that of others (Thiede, Blood 2002) suggest that FLT3 ITD mutation if present in normal karyotype patients is a key player in the pathogenesis of leukaemia and targeting this mutation may be used successfully in FLT3 ITD positive patients relapsing post HSCT. This was also the case in the two patients presented in this study. Patient UPN 952, male, 53 years old, AML M4 (FAB), normal karyotype, received alloHSCT in 2 CR after completion of two lines of remission induction therapies (first: DA3+7 (Daunorubicin, Ara-C), HAM, high dose Ara-C; second: ICE (IDA, Ara-C, VP-16)) and then promptly transplanted. He relapsed 56 days after transplantation. Patient UPN 938, female, 50 years old, AML, normal karyotype, received as an induction DA 3+7, and for consolidation HAM and high dose Ara-C and completed only one course of the maintenance therapy and transplanted in CR. Both cases were transplanted from unrelated donors (10/10 HLA A, B, C, DR and DQ alleles matched), they received myeloablative conditioning (i.v. Busulfan and Cyclophosphamide) and Cyclosporin A as GvHD prophylaxis. At relapse they received salvage chemotherapy tailored to their biological performance (UPN 938 having Age Adjusted Charlson Comorbidity Index 3 received FLAG and UPN 952 with the index 9, ECOG 3, DA2+5). Both cases received in addition Sorafenib (two times 400 mg per day). The response was prompt and the marrow was free from blasts beginning from 11 day post chemotherapy. UPN 952 received as a maintenance only one course of 6-TG with low dose of Ara-C. Due to the substantial comorbidity and liver toxicity WHO3 further chemotherapy treatment was terminated. The patient was left only on Sorafenib (2 times 200 mg per day). UPN 938 was receiving the maintenance therapy (AML protocol) in 6 – 8 weeks intervals based on low dose Ara-C with 6TG or DNR and also Sorafenib (2 times 200 mg per day). In both cases FLT3 signalling pathway (FLT3 Pathway Mutation PCR Array, SABiosciences, Qiagen) revealed a lack of any additional mutation at the check points in FLT3, KRAS, HRAS, NRAS, MEK1, PIK3CA, BRAF and PTEN genes. UPN 938 had in addition to FLT3 ITD mutation c.1807_1808insATGAATATGATCTCAAAT (p.K602_W603insYEYDLK) insertion which relevance was not so far describe. Sorafenib resistance mutations were not found. The patients were on Sorafenib for 8 (UPN 938) and 9 months (UPN 952). The course of UPN938 was uncomplicated. The second case showed mild aGvHD symptoms evolving into cGvHD (skin lesions and dry eye) slowed down with rapamicine. Up to now the patients are in CR and free from FLT3 ITD. The main observation points: - Sorafenib with chemotherapy tailored to the biologic performance of patients contributes to the efficient salvage in patients with relapsing FLT3 ITD positive AML post HSCT and is also effective given alone as a maintenance. - Side effects were seen in one out of two patients likely associated with the blocking of VEGFR signalling transmission (hand foot skin rash, von Willebrandt factor activity – 388%). Conclusion: The use of multikinase inhibitor (Sorafenib) contributes effectively to salvage therapy in FLT3+ AML patients relapsing post alloHSCTSorafenib given alone is able to maintain long-lasting remissionSide effects are individually dependent Supported by NBiR CellsTherpy grant and Byer Health Care Disclosures No relevant conflicts of interest to declare.

Background Surgery for pain in the right lower quadrant of the abdomen remains a clinical dilemma. This prospective study assessed the accuracy of preoperative unenhanced helical computed tomography (CT) in the evaluation of patients with suspected acute appendicitis. Methods One hundred and three adult patients with suspected acute appendicitis underwent unenhanced helical CT of the abdomen. Subsequently, all patients underwent laparoscopic inspection of the abdominal cavity by a surgeon who was blinded to the diagnosis suggested by CT. Patients underwent appropriate surgical therapy accordingly. Follow-up was at least 6 weeks. Results Appendicitis was diagnosed by CT in 83 patients (80·6 per cent). Acute appendicitis was identified during laparoscopy in 87 patients (84·5 per cent). Prospective interpretation of CT images yielded a sensitivity of 95·4 per cent and a specificity of 100 per cent for the diagnosis of acute appendicitis. There were four false-negative scans. In 12 of 20 patients without signs of appendicitis on CT, the scan established the presence of other pathology. At operation no additional pathology was observed in this group and all other diagnoses proved to be correct. Conclusion Plain helical CT in patients with suspected acute appendicitis provides an accurate diagnosis without the disadvantages of contrast enhancement.

Thorough knowledge of anatomy and anatomical variations are prerequisites for liver transplantation. In whole organ liver transplantation, knowledge of anatomy of hilar structures and hepatic veins is necessary but in split liver transplantation additional insight in intrahepatic anatomy is of paramount importance to ensure two viable grafts of sufficient size and with usable vascular and biliary tributaries. Understanding of the concept of functional anatomy and segmentation of the liver represents the basis for splitting the liver for transplantation purposes. In this chapter both the concept of functional anatomy of the liver and the descriptive anatomy of the hepatoduodenal ligament, the intraparenchymal branches of the portal triads and hepatic veins are pointed out. Both aspects are covered with splitting the liver for transplantation in mind. However, the next chapter focuses entirely on the consequences of liver anatomy for the splitting surgeon.

Background: A randomized clinical trial was performed to compare open appendectomy (OA) and laparoscopic appendectomy (LA). Methods: 201 patients with similar characteristics of appendicitis were randomized to either OA or LA. Operative time and technique, reintroduction of diet, postoperative pain, use of analgesia, hospital stay, and complications were documented. Results: 104 patients were allocated to the OA group and 97 to the LA group. Postoperative pain was significantly less in the LA group on the 1st (p < 0.001) and 2nd (p < 0.001) postoperative day, resulting in less use of analgesics on both days (p < 0.001). Restoration of diet was similar in both groups. Mean operative time was longer in the LA group: 61 vs 41 min (p < 0.001). Postoperative complications did not differ in either group, except for wound infections (six OA group vs zero LA group, p < 0.05). Mean hospital stay was similar in both groups. Conclusions: LA results in less postoperative pain and fewer wound infections. The laparoscopic procedure is technically more demanding to perform, resulting in longer operative time.

Abstract 4487 HLA mismatch constitute the risk of aGvHD. However, environmental factors play a significant role what is exemplified by the protective effect of germ free environment. This association can be discussed on the ground of a significant role of cytokine milieu which determines lymphocytes subsets differentiation. Our previous work showed a decrease of proportions of Th17 cells in blood at the onset of aGvHD (Dlubek et al., Transplantation Proceedings 2010; 42(8):3277-9). These cells, however, were seen at the tissue site likely being marginalized during aGvHD process. In this study we looked at Th17 lymphocytes proportions in blood of patients post alloHSCT in the context of an increase of C-reactive protein in serum and NOD2/CARD15 gene mutations. Eighty four patients (median age: 41 yrs, range: 1–64 yrs) entered the study. All of them were clinically followed for the presence of infections complications and aGvHD. Blood work was done at the very first day of the later complications and in addition routinely in one week intervals beginning from the day of transplantation. The cells were labeled for CD4 (Becton Dickinson, San Jose, CA, USA), intracellular IL-17A (e-biosciences, San Diego, CA, U04)SA) and FoxP3 (Becton Dickinson, San Jose, CA, USA) detection. CRP was measured in serum using a Behring BN Prospect nephelometer (Dade Behring Inc., Marburg, Germany). All studied cases were genotyped for NOD2/CARD15 mutations: 104C-T (SNP8, Arg702W; rs.2066844), 2722G-C (SNP12, Gly908Arg; rs.2066845), and 3020insC (SNP13, Leu1007fsinsC; rs.2066847) with the use of RFLP-PCR technique. It was found: Acute GvHD manifestation was seen in 34 (40%) patients among them 22 (26%) had only skin symptoms and 12 (14%) presented gut manifestation. Gut aGvHD was seen at later time post HSCT than skin aGvHD (median 33 vs. 21 days, M-W U test p=0.035).Twelve patients had NOD2/CARD15 mutations. They had lower proportions of Th17 lymphocytes in blood as compared to those lacking these mutations (median 0.03 vs. 0.07%, M-W U test p=0.052) and 6 (50%) of them presented aGvHD.CRP levels measured from 3 to 5 days before clinically apparent aGvHD were significantly higher in patients with gut symptoms as compared to those presented only skin lesions (median 66.8 vs. 7.61 mg/L, M-W U test p=0.020). Notably, patients having only skin symptoms had rather lower CRP levels in sera than those without aGvHD (median 7.61 vs. 27.6 mg/L, M-W U test p=0.028).In patients with gut manifestation FoxP3+CD4+ cells proportions were negatively correlated with serum CRP levels (Spearman r −0.664, p=0.018) what was not seen in patients presented only skin symptoms aGvHD (0.068, p=0.769). In conclusion: CRP elevation constituted the risk of gut aGvHD and was associated with rather exhausted T regulatory cell function. NOD2/CARD15 mutations associated factors contributed to the lowering of Th17 cells in blood what was previously found to be associated with the overt manifestation of aGvHD (Dlubek et al., Transplantation Proceedings 2010; 42(8):3277-9). These both findings are in line with the hypothetical role of microbial pathogens which via inflammation favor differentiation of lymphocytes to Th17 positive cell subset. Disclosures: No relevant conflicts of interest to declare.

Abstract 4552 It is known that CMV infection/reactivation influences the survival of patients post HSCT. Risk factors are associated with both (i) the CMV serostatus of donors and recipients and (ii) the immunological potential of the recipients post transplantation. In this study 142 patients - 69 males and 73 females suffering from hematological malignancies, SAA and inborn errors at the age from 0.6 to 64 (median 40) - were followed for the presence of CMV and EBV DNA copies and the presence of CMV and EBV IgM antibodies in blood post HSCT (61 sibling and 81 matched unrelated donors) with 625 days of median observation time. In addition the number of lymphocytes in blood and the proportions and numbers of CD4, CD8, CD20 positive cells in blood were analyzed in association with the CMV and EBV findings. CMV and EBV DNA copies were determined with the use of qPCR at 1-week intervals until 30 day post HSCT, then monthly until one year post HSCT and then at routine follow-up examinations (usually in 3 months intervals), and always when clinically suggested. Values exceeding 50 copies/10^5 cells were recognized as positive and clinically significant. At the same time the presence of IgM and IgG antibodies specific for CMV and EBV were examined with the use of ELISA. The first positive results for CMV DNA copies were seen at median time of 48 days post-transplant and IgM antibodies if present were detected in a majority of cases within 30 days after DNA copies finding. EBV copies were detected at the similar time post transplantation (median day of positive results: 48 days) and IgM antibodies if present were usually seen within 2 months after DNA copies detection. Notably, in 9 patients EBV reactivations were associated by the appearance of IgM CMV but not EBV antibodies. Therefore the presence of IgM antibodies against either CMV or EBV was recognized as a sign of the ability of a given patient to respond immunologically to CMV or EBV viruses. For the sake of this study the patients group was divided into subgroups as follow: (1) CMV and/or EBV copies present but IgM antibodies absent, (2) CMV and/or EBV DNA copies as well as CMV and/or EBV antibodies present, (3) neither CMV or EBV copies nor IgM antibodies present, (4) CMV and EBV DNA copies absent but IgM CMV and/or EBV antibodies present. Patients lacking DNA copies or having both copies and IgM antibodies against CMV and/or EBV had significantly better survival than those having DNA copies but lacking IgM antibodies any time during observation period (2 yrs survival 70% vs 41%, p To understand the immunological background to this observation lymphocytosis and blood lymphocyte subsets were analyzed at 30 day post HSCT. It became apparent that pts having CMV and/or EBV copies but lacking IgM antibodies had significantly lower number of lymphocytes and their (CD4+ cells subset) in blood as compared to patients having IgM antibodies against CMV and/or EBV irrespective whether they had CMV and/or EBV DNA copies with median values as follow: 650/ul (128/ul) vs 370/ul (60/ul), p=0.020 (p=0.033) or lacked CMV and EBV DNA copies, LYM (CD4+) median values: 750/ul (161/ul) vs 370/ul (60/ul), p=0.047 (p=0.060). CMV and EBV infection/reactivation affects the survival of patients post HSCT if they are unable to generate IgM antibodies against these viruses. Inability to mount humoral response to CMV and EBV was associated with an impairment of the immune system potential (low number of lymphocytes and CD4+ cells in blood). Supported by the grants N R13 0082 06 from the Polish Ministry of Science & Higher Education. Disclosures: No relevant conflicts of interest to declare.

Abstract 4555 It is known that the pace of immunological recovery influences the outcome of HSCT and herpes viruses reactivation contributes significantly to the post transplant mortality. The aim of this study was to investigate reconstitution of subpopulations of CD4+ lymphocytes in the context of β-herpes viruses reactivation shortly post HSCT. There were two observation points: (1) at the beginning of lymphocyte recovery (number of lymphocytes > 200 cells/ul) which was seen from 9 to 31 day post HSCT median 14 and (2) 3 weeks later (median day 35 range(31-53)). CD4+ T cell subsets were defined as follow: Naive cells (N) CD4+CCR7+CD45RA+, Central memory (CM) CD4+CCR7+CD45-, Effector memory(EM) CD4+CCR7-CD45+, Terminally differentiated memory cells (TEMRA) CD4+CCR7-CD45RA+ . CMV, EBV and HHV-6 DNA copies were detected in blood with the use of QT-PCR in one week intervals. The study included 44 patients (age 1 to 64 median 45yr., 21 ric/mac, 21/23, sib/mud 21/23 mud, BM/PBPC 4/40. It was found; 1. Fifteen, 13 and 4 patients were positive for EBV, HHV-6 and CMV DNA copies during the observation period. (i) CD4+lymphocyte count was lower in patients having CMV copies as compared to those having EBV and/or HHV6 copies (both lymphocytotropic viruses) at the beginning of lymphocytes recovery (45/ul vs 120/ul p=0,047 Mann-Whitney U test), similarly behaved EM cells. 22/ul vs 84/ul EM cells (p= 0,088 Mann-Whitney U test) were seen in pts having CMV reactivation and in those with EBV and/or HHV-6 reactivation, respectively. (ii) The situation changed three weeks later, it appeared that patients having HHV6 reactivation any time during the observation period had lower CD4+ lymphocyte numbers as compared to patients lacking β-herpes viruses reactivations (CD4+ cells median 93/ul vs190/ul; p=0,096 Mann-Whitney U test) and to those with CMV and EBV reactivation. Again EM cells contributed greatly to these differences. EM CD4+ lymphocytes were 41/ul, 105/ul, 80/ul and 160/ul in patients having HHV-6, CMV or EBV and lacking beta-herpes viruses reactivation, respectively. EM CD4+ lymphocyte numbers for HHV6+ patients as compared to patients lacking β-herpes viruses reactivations was 41/ul vs155/ul.(p=0,012 Mann-Whitney U test). Conclusions: It appears that HHV6 is the main player in shaping lymphocyte reconstitution post transplant affecting predominantly EM cells what influences the outcome of transplantation. In this group of patients probability of one year survival was 50% vs 65%, in patients having and lacking HHV6, respectively. Disclosures: No relevant conflicts of interest to declare.

IL-17 is involved in chronic inflammation and autoimmunity, this cytokine producing cells are well characterized in the mouse model. In man, situation is more complex due to the genetic variability, co-morbidities and environmental factors influencing the immune response. In patients post HSCT alloreactivity with the presence of aGvHD is a major factor affecting the outcome of transplantation. Understanding of the role of cells involved in regulation of the immune system is crucial for the treatment tailoring to favour tolerance but not making the recipient more prone to opportunistic infections and leukaemia relapse. In this study 27 patients post HSCT were investigated for the presence of regulatory cells and those producing IFNgamma and IL-17 in blood. Blood was collected at the time of haematological recovery or when the first clinical symptoms of aGvHD became apparent and then in one week intervals until +60 day post transplantation. PBMC were stimulated for 4 hrs with BD Leukocyte Activation Cocktail in the presence of Golgi Stop (BD, Erembodegen, Belgium) and then stained extracellularly with anti-CD4, permeabilized with Fixation/Permeabilization Concentrate and Diluent (eBioscience, San Diego, CA) and finally stained with anti-IFNgamma (BD), anti-IL-17 (eBioscience) anti FoxP3 (eBioscience). CD4+ cells subpopulations were analysed according to the expression of the stained features. It was found: FoxP3+CD4+ cells were in higher proportions in pts with aGvHD (results from all time-points taken together) (9.93%±0.61 vs 8.2%±0.50, n=98, p=0.040, U Mann-Whitney test) IFNgamma producing CD4+ lymphocytes were in higher proportions (0,34 vs 0.14, ns) in blood samples taken from patients lacking as compared to those having aGvHD. CD4+IL-17+ lymphocytes proportions increased from 1.39%±0.42 to 5.33%±2.45 (p=0.04, Wilcoxon Test for pairs) one week before aGvHD. Notably, at the time of full blown aGvHD the proportions of CD4+IL-17+ cells were lower as compared to the results of previous measurements (0.74%±0.29, p=0.008, Wilcoxon Test for pairs). Taking all results together the proportion of CD4+IL17+ lymphocytes were lower in patients having as compared to those lacking aGVHD (0.93%±0.27 vs 1,53%±0,41, p=0.05, U Mann-Whitney test). It appears that: FoxP3 positive cells expand during aGvHD likely as a response to alloreactive stimulation. INFgamma+ CD4+ cells benefit the course post HSCT making the patients less susceptible to aGvHD. CD4+IL17+ cells are likely involved at the early stage of aGvHD patho-mechanism heralding the clinical manifestation of this complication, but then they disappear from blood, possibly being marginalized in the inflamed tissues.

Cytokines belonging to interleukin -17 family (IL-17A, B, C, D, E and F) are produced mainly by activated and memory CD4+ cells which are described as Th17. These cells (IL-17+CD4+) promote autoimmune reaction (mouse model), were found in tissues involved in autoimmune process in man and produce IFNgamma. Therefore, Th17 cells constitute a population which may contribute to aGvHD together with Treg cells (FoxP3+). In this study we examined the cytoplasmic expression of IL-17A and FoxP3 in CD4+ lymphocytes in 6 pts post HSCT (3 MUD and 3 SIB, median (mean) age: 35 (33,9) years, 3 AML, 2 ALL-T, 1 SCID). Three of them developed aGvHD. All consecutive pts were included to the study. Peripheral blood lymphocytes were examined or at the very first day of aGvHD manifestation either at the beginning of hematological reconstitution and thereafter in one week intervals. Nine healthy individuals served as controls. PBMC were stimulated with BD Leukocyte Activation Cocktail containing mixture of PMA, Ionomycin and brefeldin A in the presence of BD GolgiStop then stained with CD4, IL-17A and FoxP3 monoclonal antibodies and analyzed in CD4+ and CD4- lymphocytes gates by three colour flow cytometry. Therefore, all results reflect the situation in stimulated cells. FoxP3 mRNA was measured in unstimulated PBMC with the use of qPCR. We found: Patients post HSCT (during 6 weeks period post transplant) had higher percentages of IL-17A+ cells in CD4+ lymphocytes as compared to healthy controls (2.0%±1.08 vs 0.2%±0.06, p=0.006) independently whether they developed or not aGvHD. In contrast to the lower proportion of total CD4+ cells in pts post HSCT then in controls (18.5%±2.1 vs 34.9%±0.7, p=0.0006) proportions of a subset of CD4+ lymphocytes which produced IL-17A+ were rather higher in pts post HSCT as compared to controls (0.2%±0.06 vs 0.07%±0.02, ns). IL-17A+ cells contributed rather to CD4+ then CD4- lymphocytes in controls (0.2%±0.06 vs 0.05%±0.01, p=0.007) as well as in pts post HSCT (2.0%±1.08 vs 0.05%±0.01, p=0.00004) FoxP3 positive cells in stimulated CD4+ lymphocytes were in lower proportions in pts post HSCT then in healthy individuals (51.6%±3.9 vs 62.2%±2.9, ns). Percentages of FoxP3+CD4+ in stimulated lymphocytes (cytometry) correlated with quantities of FoxP3 mRNA in unstimulated PBMC (qPCR) (R=0,42, p=0,065). Notably, there was a negative correlation between proportions of FoxP3 and IL-17A positive cells in CD4+ lymphocytes in patients post HSCT (R= − 0,55; p=0,012). In conclusion, HSCT was associated with: an increase in proportion of Th17 cells, a higher contribution of IL-17A+ cells to the pool of CD4+ cells as compared to healthy individuals and a network in the immune system in which CD4+ FoxP3 + cells and Th17 cells play an opposite role.

Allogeneic haematopoietic stem cell transplantation (HSCT) has relied on myeloablative conditioning (MAC) to ablate malignancy and control graft rejection. Conditioning is vital for HSC engraftment, however it up-regulates inflammatory cytokine and endotoxin levels, activates dendritic cells (DC) and initiates GVHD. Non-myeloablative, but immunosuppressive, reduced intensity conditioning (RIC) is increasingly used to combat treatment related mortality (TRM) associated with MAC. RIC reduces TRM and delays GVHD onset, but overall GVHD incidence is unchanged. We hypothesise that this is due to persistence of host DC. No clinically relevant in vivo murine models of RIC allogeneic HSCT have been reported to date, preventing studies of the mechanisms of RIC-associated delayed-onset GVHD. We developed two murine models of pre-transplant conditioning, which result in GVHD after infusion of donor cells [either MHC mismatched (UBI-GFP/BL6→BALB/c) or MHC-matched minor antigen mismatch (miHA; UBI-GFP/BL6→BALB.B), the latter mimicking allogeneic HLA-identical sibling HSCT]. These models combine clinical doses of cytotoxics (cyclophosphamide and fludarabine) with radiation, enabling us to monitor effects of these regimens on DC subsets and activation daily during conditioning and thereafter. GVHD onset is significantly delayed after RIC compared to MAC in HSCT recipients in MHC mismatched (MAC d+6 vs. RIC d+18–35; p These findings suggest that the major factors contributing to delayed-onset GVHD post-HSCT in RIC recipients are persistence of host DC and delayed emergence of activated donor DC. Delineating the effect of MAC and RIC regimens on DC provides a rationale to target activated DC as GVHD therapy. Fig 1. Delayed onset GvHD after RIC. BALB/c or BALB B mice received RIC or MAC followed by transplantation with UBI-GFP/BL6 BM and spleen cells. Mice were scored for GvHD and euthanased when they met ethical criteria for sacrifice. Fig 1. Delayed onset GvHD after RIC. BALB/c or BALB B mice received RIC or MAC followed by transplantation with UBI-GFP/BL6 BM and spleen cells. Mice were scored for GvHD and euthanased when they met ethical criteria for sacrifice. Fig 2. Comparison of host and donor chimerism in MHC mismatch HSCT recipients. A) Percent total chimerism in RIC and MAC recipients at time of total GvHD B) Percent chimerism of activated BM DC at time of controlled sacrifice C) Percent chimerism of activated BM DC at time of total GvHD. Fig 2. Comparison of host and donor chimerism in MHC mismatch HSCT recipients. A) Percent total chimerism in RIC and MAC recipients at time of total GvHD B) Percent chimerism of activated BM DC at time of controlled sacrifice C) Percent chimerism of activated BM DC at time of total GvHD.

Graft versus Host Disease (GVHD) and treatment related mortality (TRM) are the major limitations to the widespread application of allogeneic haematopoietic stem cell transplantation (HSCT) for haematological and non-haematological malignancies. Dendritic cells (DC) as the key initiators and directors of the immune response are central to allogeneic transplant interactions. Preparative conventional conditioning (CC) regimens aim to control disease and ablate the host immune response to facilitate normal haematopoietic reconstitution. The conditioning also unleashes a cytokine storm that activates the residual host immune system, driving host DC and donor T cell interactions that result in GVHD. Reducing the intensity of conditioning (RIC) regimens maintains immune anti-leukaemic activity of T replete HSCT, reduces TRM and delays the onset of GVHD, but the overall incidence of GVHD is unchanged. We hypothesise that this is due to increased persistence of host DC. We propose that strategic administration of DC depleting antibodies could be an effective means to control GVHD. Whilst there is some information on the effects of CC on DC; we have shown that mature plasmacytoid (p) DC are increased in mouse spleen after conditioning by radiation, there is no information on the effects of RIC on DC. We have established murine models of conditioning (CC = Cyclophosphamide [CY] + 800cGy and RIC = Fludarabine [FLU] + CY + 200cGy). The effects on DC numbers, activation and subset composition (myeloid (m) DC and DC), cytokine and systemic endotoxin levels were studied on each day of the conditioning regimens in the absence of HSCT. Mice receiving CC have a significantly higher percentage of DC which are pDC compared to RIC (p

The BM contains progenitor cells that give rise to hematopoietic tissue and also more primitive mesenchymal stem cells (MSC) which may differentiate into other tissues including endothelial cells. This potential of BM cells has already been employed in clinical studies suggesting that implantation of autologous BM cells may induce angiogenesis in ischemic tissues. In the present study 10 male pts with critical leg ischemia (41–64 yrs) suffering from pain at rest and/or foot ischemic ulceration (9/10 pts) with ABI (ankle/brachial index)