Xyloglucan deficiency leads to a reduction in turgor pressure and changes in cell wall properties, affecting early seedling establishment.

Xyloglucan is believed to play a significant role in cell wall mechanics of dicot plants. Surprisingly, Arabidopsis plants defective in xyloglucan biosynthesis exhibit nearly normal growth and development. We investigated a mutant line, cslc-Δ5, lacking activity in all five Arabidopsis cellulose synthase like-C (CSLC) genes responsible for xyloglucan backbone biosynthesis. We observed that this xyloglucan-deficient line exhibited reduced cellulose crystallinity and increased pectin levels, suggesting the existence of feedback mechanisms that regulate wall composition to compensate for the absence of xyloglucan. These alterations in cell wall composition in the xyloglucan-absent plants were further linked to a decrease in cell wall elastic modulus and rupture stress, as observed through atomic force microscopy (AFM) and extensometer-based techniques. This raised questions about how plants with such modified cell wall properties can maintain normal growth. Our investigation revealed two key factors contributing to this phenomenon. First, measurements of turgor pressure, a primary driver of plant growth, revealed that cslc-Δ5 plants have reduced turgor, preventing the compromised walls from bursting while still allowing growth to occur. Second, we discovered the conservation of elastic asymmetry (ratio of axial to transverse wall elasticity) in the mutant, suggesting an additional mechanism contributing to the maintenance of normal growth. This novel feedback mechanism between cell wall composition and mechanical properties, coupled with turgor pressure regulation, plays a central role in the control of plant growth and is critical for seedling establishment in a mechanically challenging environment by affecting shoot emergence and root penetration.

Synthesis and import of GDP-l-fucose into the Golgi affect plant-water relations.

Land plants evolved multiple adaptations to restrict transpiration. However, the underlying molecular mechanisms are not sufficiently understood. We used an ozone-sensitivity forward genetics approach to identify Arabidopsis thaliana mutants impaired in gas exchange regulation. High water loss from detached leaves and impaired decrease of leaf conductance in response to multiple stomata-closing stimuli were identified in a mutant of MURUS1 (MUR1), an enzyme required for GDP-l-fucose biosynthesis. High water loss observed in mur1 was independent from stomatal movements and instead could be linked to metabolic defects. Plants defective in import of GDP-l-Fuc into the Golgi apparatus phenocopied the high water loss of mur1 mutants, linking this phenotype to Golgi-localized fucosylation events. However, impaired fucosylation of xyloglucan, N-linked glycans, and arabinogalactan proteins did not explain the aberrant water loss of mur1 mutants. Partial reversion of mur1 water loss phenotype by borate supplementation and high water loss observed in boron uptake mutants link mur1 gas exchange phenotypes to pleiotropic consequences of l-fucose and boron deficiency, which in turn affect mechanical and morphological properties of stomatal complexes and whole-plant physiology. Our work emphasizes the impact of fucose metabolism and boron uptake on plant-water relations.

Stiffness transitions in new walls post-cell division differ between Marchantia polymorpha gemmae and Arabidopsis thaliana leaves.

Plant morphogenesis is governed by the mechanics of the cell wall-a stiff and thin polymeric box that encloses the cells. The cell wall is a highly dynamic composite material. New cell walls are added during cell division. As the cells continue to grow, the properties of cell walls are modulated to undergo significant changes in shape and size without breakage. Spatial and temporal variations in cell wall mechanical properties have been observed. However, how they relate to cell division remains an outstanding question. Here, we combine time-lapse imaging with local mechanical measurements via atomic force microscopy to systematically map the cell wall's age and growth, with their stiffness. We make use of two systems, Marchantia polymorpha gemmae, and Arabidopsis thaliana leaves. We first characterize the growth and cell division of M. polymorpha gemmae. We then demonstrate that cell division in M. polymorpha gemmae results in the generation of a temporary stiffer and slower-growing new wall. In contrast, this transient phenomenon is absent in A. thaliana leaves. We provide evidence that this different temporal behavior has a direct impact on the local cell geometry via changes in the junction angle. These results are expected to pave the way for developing more realistic plant morphogenetic models and to advance the study into the impact of cell division on tissue growth.

hENT1 Predicts Benefit from Gemcitabine in Pancreatic Cancer but Only with Low CDA mRNA.

Gemcitabine or 5-fluorouracil (5-FU) based treatments can be selected for pancreatic cancer. Equilibrative nucleoside transporter 1 (hENT1) predicts adjuvant gemcitabine treatment benefit over 5-FU. Cytidine deaminase (CDA), inside or outside of the cancer cell, will deaminate gemcitabine, altering transporter affinity. ESPAC-3(v2) was a pancreatic cancer trial comparing adjuvant gemcitabine and 5-FU. Tissue microarray sections underwent in situ hybridization and immunohistochemistry. Analysis of both CDA and hENT1 was possible with 277 patients. The transcript did not correlate with protein levels for either marker. High hENT1 protein was prognostic with gemcitabine; median overall survival was 26.0 v 16.8 months (p = 0.006). Low CDA transcript was prognostic regardless of arm; 24.8 v 21.2 months with gemcitabine (p = 0.02) and 26.4 v 14.6 months with 5-FU (p = 0.02). Patients with low hENT1 protein did better with 5-FU, but only if the CDA transcript was low (median survival of 5-FU v gemcitabine; 29.3 v 18.3 months, compared with 14.2 v 14.6 with high CDA). CDA mRNA is an independent prognostic biomarker. When added to hENT1 protein status, it may also provide treatment-specific predictive information and, within the frame of a personalized treatment strategy, guide to either gemcitabine or 5FU for the individual patient.

KATANIN and CLASP function at different spatial scales to mediate microtubule response to mechanical stress in Arabidopsis cotyledons.

Mechanical stress influences cell- and tissue-scale processes across all kingdoms. It remains challenging to delineate how mechanical stress, originating at these different length scales, impacts cell and tissue form. We combine growth tracking of cells, quantitative image analysis, as well as molecular and mechanical perturbations to address this problem in pavement cells of Arabidopsis thaliana cotyledon tissue. We show that microtubule organization based on chemical signals and cell-shape-derived mechanical stress varies during early stages of pavement cell development and is mediated by the evolutionary conserved proteins, KATANIN and CLASP. However, we find that these proteins regulate microtubule organization in response to tissue-scale mechanical stress to different extents in the cotyledon epidermis. Our results further demonstrate that regulation of cotyledon form is uncoupled from the mechanical-stress-dependent control of pavement cell shape that relies on microtubule organization governed by subcellular mechanical stress.

Nuclear calcium signatures are associated with root development.

In plants, nuclear Ca2+ releases are essential to the establishment of nitrogen-fixing and phosphate-delivering arbuscular mycorrhizal endosymbioses. In the legume Medicago truncatula, these nuclear Ca2+ signals are generated by a complex of nuclear membrane-localised ion channels including the DOES NOT MAKE INFECTIONS 1 (DMI1) and the cyclic nucleotide-gated channels (CNGC) 15s. DMI1 and CNCG15s are conserved among land plants, suggesting roles for nuclear Ca2+ signalling that extend beyond symbioses. Here we show that nuclear Ca2+ signalling initiates in the nucleus of Arabidopsis root cells and that these signals are correlated with primary root development, including meristem development and auxin homeostasis. In addition, we demonstrate that altering genetically AtDMI1 is sufficient to modulate the nuclear Ca2+ signatures, and primary root development. This finding supports the postulate that stimulus-specific information can be encoded in the frequency and duration of a Ca2+ signal and thereby regulate cellular function.

The Impact of Positive Resection Margins on Survival and Recurrence Following Resection and Adjuvant Chemotherapy for Pancreatic Ductal Adenocarcinoma.

OBJECTIVE AND BACKGROUND: Local and distant disease recurrence are frequently observed following pancreatic cancer resection, but an improved understanding of resection margin assessment is required to aid tailored therapies. METHODS: Analyses were carried out to assess the association between clinical characteristics and margin involvement as well as the effects of individual margin involvement on site of recurrence and overall and recurrence-free survival using individual patient data from the European Study Group for Pancreatic Cancer (ESPAC)-3 randomized controlled trial. RESULTS: There were 1151 patients, of whom 505 (43.9%) had an R1 resection. The median and 95% confidence interval (CI) overall survival was 24.9 (22.9-27.2) months for 646 (56.1%) patients with resection margin negative (R0 >1 mm) tumors, 25.4 (21.6-30.4) months for 146 (12.7%) patients with R1<1 mm positive resection margins, and 18.7 (17.2-21.1) months for 359 (31.2%) patients with R1-direct positive margins (P < 0.001). In multivariable analysis, overall R1-direct tumor margins, poor tumor differentiation, positive lymph node status, WHO performance status ≥1, maximum tumor size, and R1-direct posterior resection margin were all independently significantly associated with reduced overall and recurrence-free survival. Competing risks analysis showed that overall R1-direct positive resection margin status, positive lymph node status, WHO performance status 1, and R1-direct positive superior mesenteric/medial margin resection status were all significantly associated with local recurrence. CONCLUSIONS: R1-direct resections were associated with significantly reduced overall and recurrence-free survival following pancreatic cancer resection. Resection margin involvement was also associated with an increased risk for local recurrence.

PET-PANC: multicentre prospective diagnostic accuracy and health economic analysis study of the impact of combined modality 18fluorine-2-fluoro-2-deoxy-d-glucose positron emission tomography with computed tomography scanning in the diagnosis and management of pancreatic cancer.

BACKGROUND: Pancreatic cancer diagnosis and staging can be difficult in 10-20% of patients. Positron emission tomography (PET)/computed tomography (CT) adds precise anatomical localisation to functional data. The use of PET/CT may add further value to the diagnosis and staging of pancreatic cancer. OBJECTIVE: To determine the incremental diagnostic accuracy and impact of PET/CT in addition to standard diagnostic work-up in patients with suspected pancreatic cancer. DESIGN: A multicentre prospective diagnostic accuracy and clinical value study of PET/CT in suspected pancreatic malignancy. PARTICIPANTS: Patients with suspected pancreatic malignancy. INTERVENTIONS: All patients to undergo PET/CT following standard diagnostic work-up. MAIN OUTCOME MEASURES: The primary outcome was the incremental diagnostic value of PET/CT in addition to standard diagnostic work-up with multidetector computed tomography (MDCT). Secondary outcomes were (1) changes in patients' diagnosis, staging and management as a result of PET/CT; (2) changes in the costs and effectiveness of patient management as a result of PET/CT; (3) the incremental diagnostic value of PET/CT in chronic pancreatitis; (4) the identification of groups of patients who would benefit most from PET/CT; and (5) the incremental diagnostic value of PET/CT in other pancreatic tumours. RESULTS: Between 2011 and 2013, 589 patients with suspected pancreatic cancer underwent MDCT and PET/CT, with 550 patients having complete data and in-range PET/CT. Sensitivity and specificity for the diagnosis of pancreatic cancer were 88.5% and 70.6%, respectively, for MDCT and 92.7% and 75.8%, respectively, for PET/CT. The maximum standardised uptake value (SUVmax.) for a pancreatic cancer diagnosis was 7.5. PET/CT demonstrated a significant improvement in relative sensitivity (p = 0.01) and specificity (p = 0.023) compared with MDCT. Incremental likelihood ratios demonstrated that PET/CT significantly improved diagnostic accuracy in all scenarios (p < 0.0002). PET/CT correctly changed the staging of pancreatic cancer in 56 patients (p = 0.001). PET/CT influenced management in 250 (45%) patients. PET/CT stopped resection in 58 (20%) patients who were due to have surgery. The benefit of PET/CT was limited in patients with chronic pancreatitis or other pancreatic tumours. PET/CT was associated with a gain in quality-adjusted life-years of 0.0157 (95% confidence interval -0.0101 to 0.0430). In the base-case model PET/CT was seen to dominate MDCT alone and is thus highly likely to be cost-effective for the UK NHS. PET/CT was seen to be most cost-effective for the subgroup of patients with suspected pancreatic cancer who were thought to be resectable. CONCLUSION: PET/CT provided a significant incremental diagnostic benefit in the diagnosis of pancreatic cancer and significantly influenced the staging and management of patients. PET/CT had limited utility in chronic pancreatitis and other pancreatic tumours. PET/CT is likely to be cost-effective at current reimbursement rates for PET/CT to the UK NHS. This was not a randomised controlled trial and therefore we do not have any information from patients who would have undergone MDCT only for comparison. In addition, there were issues in estimating costs for PET/CT. Future work should evaluate the role of PET/CT in intraductal papillary mucinous neoplasm and prognosis and response to therapy in patients with pancreatic cancer. STUDY REGISTRATION: Current Controlled Trials ISRCTN73852054 and UKCRN 8166. FUNDING: The National Institute for Health Research Health Technology Assessment programme.

Minimally invasive and endoscopic versus open necrosectomy for necrotising pancreatitis: a pooled analysis of individual data for 1980 patients.

OBJECTIVE: Minimally invasive surgical necrosectomy and endoscopic necrosectomy, compared with open necrosectomy, might improve outcomes in necrotising pancreatitis, especially in critically ill patients. Evidence from large comparative studies is lacking. DESIGN: We combined original and newly collected data from 15 published and unpublished patient cohorts (51 hospitals; 8 countries) on pancreatic necrosectomy for necrotising pancreatitis. Death rates were compared in patients undergoing open necrosectomy versus minimally invasive surgical or endoscopic necrosectomy. To adjust for confounding and to study effect modification by clinical severity, we performed two types of analyses: logistic multivariable regression and propensity score matching with stratification according to predicted risk of death at baseline (low: <5%; intermediate: ≥5% to <15%; high: ≥15% to <35%; and very high: ≥35%). RESULTS: Among 1980 patients with necrotising pancreatitis, 1167 underwent open necrosectomy and 813 underwent minimally invasive surgical (n=467) or endoscopic (n=346) necrosectomy. There was a lower risk of death for minimally invasive surgical necrosectomy (OR, 0.53; 95% CI 0.34 to 0.84; p=0.006) and endoscopic necrosectomy (OR, 0.20; 95% CI 0.06 to 0.63; p=0.006). After propensity score matching with risk stratification, minimally invasive surgical necrosectomy remained associated with a lower risk of death than open necrosectomy in the very high-risk group (42/111 vs 59/111; risk ratio, 0.70; 95% CI 0.52 to 0.95; p=0.02). Endoscopic necrosectomy was associated with a lower risk of death than open necrosectomy in the high-risk group (3/40 vs 12/40; risk ratio, 0.27; 95% CI 0.08 to 0.88; p=0.03) and in the very high-risk group (12/57 vs 28/57; risk ratio, 0.43; 95% CI 0.24 to 0.77; p=0.005). CONCLUSION: In high-risk patients with necrotising pancreatitis, minimally invasive surgical and endoscopic necrosectomy are associated with reduced death rates compared with open necrosectomy.

Pavement cells and the topology puzzle.

D'Arcy Thompson emphasised the importance of surface tension as a potential driving force in establishing cell shape and topology within tissues. Leaf epidermal pavement cells grow into jigsaw-piece shapes, highly deviating from such classical forms. We investigate the topology of developing Arabidopsis leaves composed solely of pavement cells. Image analysis of around 50,000 cells reveals a clear and unique topological signature, deviating from previously studied epidermal tissues. This topological distribution is established early during leaf development, already before the typical pavement cell shapes emerge, with topological homeostasis maintained throughout growth and unaltered between division and maturation zones. Simulating graph models, we identify a heuristic cellular division rule that reproduces the observed topology. Our parsimonious model predicts how and when cells effectively place their division plane with respect to their neighbours. We verify the predicted dynamics through in vivo tracking of 800 mitotic events, and conclude that the distinct topology is not a direct consequence of the jigsaw piece-like shape of the cells, but rather owes itself to a strongly life history-driven process, with limited impact from cell-surface mechanics.

Stomatal Opening Involves Polar, Not Radial, Stiffening Of Guard Cells.

It has long been accepted that differential radial thickening of guard cells plays an important role in the turgor-driven shape changes required for stomatal pore opening to occur [1-4]. This textbook description derives from an original interpretation of structure rather than measurement of mechanical properties. Here we show, using atomic force microscopy, that although mature guard cells display a radial gradient of stiffness, this is not present in immature guard cells, yet young stomata show a normal opening response. Finite element modeling supports the experimental observation that radial stiffening plays a very limited role in stomatal opening. In addition, our analysis reveals an unexpected stiffening of the polar regions of the stomata complexes, both in Arabidopsis and other plants, suggesting a widespread occurrence. Combined experimental data (analysis of guard cell wall epitopes and treatment of tissue with cell wall digesting enzymes, coupled with bioassay of guard cell function) plus modeling lead us to propose that polar stiffening reflects a mechanical, pectin-based pinning down of the guard cell ends, which restricts increase of stomatal complex length during opening. This is predicted to lead to an improved response sensitivity of stomatal aperture movement with respect to change of turgor pressure. Our results provide new insight into the mechanics of stomatal function, both negating an established view of the importance of radial thickening and providing evidence for a significant role for polar stiffening. Improved stomatal performance via altered cell-wall-mediated mechanics is likely to be of evolutionary and agronomic significance.

The role of induction chemotherapy + chemoradiotherapy in localised pancreatic cancer: initial experience in Scotland.

BACKGROUND: Despite being relatively rare pancreatic cancer is one of the highest causes of death. Even within the potentially resectable group outcomes are poor. We present our initial experiences utilising a neoadjuvant approach to localised pancreatic cancer, evaluating survival, response rates and tolerability. METHODS: This was a retrospective analysis of a prospectively maintained database. Patients from 2012 to 2015 referred to a busy regional Hepato-Pancreatic Biliary (HPB) MDT were included. Patients were classified according to respectability criteria (utilising NCCN guidelines) and a treatment plan agreed. Systemic therapy with either FOLFIRINOX or Gem/Cap was delivered followed by chemoradiotherapy if disease remained localised. Toxicity, response, pathological outcomes and survival were all recorded. RESULTS: A total of 85 patients were included in the study: 45 had initially resectable disease; 19 required a response for resection and 21 had locally advanced inoperable disease; 34 patients underwent resection. The median survival for the potentially resectable group was 22.2 months while for those undergoing resection it was 37 months. CONCLUSIONS: We have demonstrated that a neoadjuvant approach is deliverable and tolerable. In addition we have demonstrated impressive survival results in patients undergoing resection with no detriment in outcome for those not proceeding to surgery.

Describing Peripancreatic Collections According to the Revised Atlanta Classification of Acute Pancreatitis: An International Interobserver Agreement Study.

OBJECTIVES: Severe acute pancreatitis is associated with peripancreatic morphologic changes as seen on imaging. Uniform communication regarding these morphologic findings is crucial for accurate diagnosis and treatment. For the original 1992 Atlanta classification, interobserver agreement is poor. We hypothesized that for the revised Atlanta classification, interobserver agreement will be better. METHODS: An international, interobserver agreement study was performed among expert and nonexpert radiologists (n = 14), surgeons (n = 15), and gastroenterologists (n = 8). Representative computed tomographies of all stages of acute pancreatitis were selected from 55 patients and were assessed according to the revised Atlanta classification. The interobserver agreement was calculated among all reviewers and subgroups, that is, expert and nonexpert reviewers; interobserver agreement was defined as poor (≤0.20), fair (0.21-0.40), moderate (0.41-0.60), good (0.61-0.80), or very good (0.81-1.00). RESULTS: Interobserver agreement among all reviewers was good (0.75 [standard deviation, 0.21]) for describing the type of acute pancreatitis and good (0.62 [standard deviation, 0.19]) for the type of peripancreatic collection. Expert radiologists showed the best and nonexpert clinicians the lowest interobserver agreement. CONCLUSIONS: Interobserver agreement was good for the revised Atlanta classification, supporting the importance for widespread adaption of this revised classification for clinical and research communications.

Pancreatic anastomosis after pancreatoduodenectomy: A position statement by the International Study Group of Pancreatic Surgery (ISGPS).

BACKGROUND: Clinically relevant postoperative pancreatic fistula (grades B and C of the ISGPS definition) remains the most troublesome complication after pancreatoduodenectomy. The approach to management of the pancreatic remnant via some form of pancreatico-enteric anastomosis determines the incidence and severity of clinically relevant postoperative pancreatic fistula. Despite numerous trials comparing diverse pancreatico-enteric anastomosis techniques and other adjunctive strategies (pancreatic duct stenting, somatostatin analogues, etc), currently, there is no clear consensus regarding the ideal method of pancreatico-enteric anastomosis. METHODS: An international panel of pancreatic surgeons working in well-known, high-volume centers reviewed the best contemporary literature concerning pancreatico-enteric anastomosis and worked to develop a position statement on pancreatic anastomosis after pancreatoduodenectomy. RESULTS: There is inherent risk assumed by creating a pancreatico-enteric anastomosis based on factors related to the gland (eg, parenchymal texture, disease pathology). None of the technical variations of pancreaticojejunal or pancreaticogastric anastomosis, such as duct-mucosa, invagination method, and binding technique, have been found to be consistently superior to another. Randomized trials and meta-analyses comparing pancreaticogastrostomy versus pancreaticojejunostomy yield conflicting results and are inherently prone to bias due to marked heterogeneity in the studies. The benefit of stenting the pancreatico-enteric anastomosis to decrease clinically relevant postoperative pancreatic fistula is not supported by high-level evidence. While controversial, somatostatin analogues appear to decrease perioperative complications but not mortality, although consistent data across the more than 20 studies addressing this topic are lacking. The Fistula Risk Score is useful for predicting postoperative pancreatic fistula as well as for comparing outcomes of pancreatico-enteric anastomosis across studies. CONCLUSION: Currently, no specific technique can eliminate development of clinically relevant postoperative pancreatic fistula. While consistent practice of any standardized technique may decrease the rate of clinically relevant postoperative pancreatic fistula, experienced surgeons can have lower postoperative pancreatic fistula rates performing a variety of techniques depending on the clinical situation. There is no clear evidence on the benefit of internal or external stenting after pancreatico-enteric anastomosis. The use of somatostatin analogues may be important in decreasing morbidity after pancreatoduodenectomy, but it remains controversial. Future studies should focus on novel approaches to decrease the rate of clinically relevant postoperative pancreatic fistula with appropriate risk adjustment.

Investigating Various Thresholds as Immunohistochemistry Cutoffs for Observer Agreement.

BACKGROUND: Clinical translation of immunohistochemistry (IHC) biomarkers requires reliable and reproducible cutoffs or thresholds for interpretation of immunostaining. Most IHC biomarker research focuses on the clinical relevance (diagnostic, prognostic, or predictive utility) of cutoffs, with less emphasis on observer agreement using these cutoffs. From the literature, we identified 3 commonly used cutoffs of 10% positive epithelial cells, 20% positive epithelial cells, and moderate to strong staining intensity (+2/+3 hereafter) to use for investigating observer agreement. MATERIALS AND METHODS: A series of 36 images of microarray cores stained for 4 different IHC biomarkers, with variable staining intensity and percentage of positive cells, was used for investigating interobserver and intraobserver agreement. Seven pathologists scored the immunostaining in each image using the 3 cutoffs for positive and negative staining. Kappa (κ) statistic was used to assess the strength of agreement for each cutoff. RESULTS: The interobserver agreement between all 7 pathologists using the 3 cutoffs was reasonably good, with mean κ scores of 0.64, 0.59, and 0.62, respectively, for 10%, 20%, and +2/+3 cutoffs. A good agreement was observed for experienced pathologists using the 10% cutoff, and their agreement was statistically higher than for junior pathologists (P=0.02). In addition, the mean intraobserver agreement for all 7 pathologists using the 3 cutoffs was reasonably good, with mean κ scores of 0.71, 0.60, and 0.73, respectively, for 10%, 20%, and +2/+3 cutoffs. For all 3 cutoffs, a positive correlation was observed with perceived ease of interpretation (P<0.003). Finally, cytoplasmic-only staining achieved higher agreement using all 3 cutoffs than mixed staining patterns. CONCLUSIONS: All 3 cutoffs investigated achieve reasonable strength of agreement, modestly decreasing interobserver and intraobserver variability in IHC interpretation. These cutoffs have previously been used in cancer pathology, and this study provides evidence that these cutoffs can be reproducible between practicing pathologists.

Auxin influx importers modulate serration along the leaf margin.

Leaf shape in Arabidopsis is modulated by patterning events in the margin that utilize a PIN-based auxin exporter/CUC2 transcription factor system to define regions of promotion and retardation of growth, leading to morphogenesis. In addition to auxin exporters, leaves also express auxin importers, notably members of the AUX1/LAX family. In contrast to their established roles in embryogenesis, lateral root and leaf initiation, the function of these transporters in leaf development is poorly understood. We report that three of these genes (AUX1, LAX1 and LAX2) show specific and dynamic patterns of expression during early leaf development in Arabidopsis, and that loss of expression of all three genes is required for observation of a phenotype in which morphogenesis (serration) is decreased. We used these expression patterns and mutant phenotypes to develop a margin-patterning model that incorporates an AUX1/LAX1/LAX2 auxin import module that influences the extent of leaf serration. Testing of this model by margin-localized expression of axr3-1 (AXR17) provides further insight into the role of auxin in leaf morphogenesis.

IP-10/CXCL10 induction in human pancreatic cancer stroma influences lymphocytes recruitment and correlates with poor survival.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by an abundant desmoplastic reaction driven by pancreatic stellate cells (PSCs) that contributes to tumor progression. Here we sought to characterize the interactions between pancreatic cancer cells (PCCs) and PSCs that affect the inflammatory and immune response in pancreatic tumors. Conditioned media from mono- and cocultures of PSCs and PCCs were assayed for expression of cytokines and growth factors. IP-10/CXCL10 was the most highly induced chemokine in coculture of PSCs and PCCs. Its expression was induced in the PSCs by PCCs. IP-10 was elevated in human PDAC specimens, and positively correlated with high stroma content. Furthermore, gene expression of IP-10 and its receptor CXCR3 were significantly associated with the intratumoral presence of regulatory T cells (Tregs). In an independent cohort of 48 patients with resectable pancreatic ductal adenocarcinoma, high IP-10 expression levels correlated with decreased median overall survival. Finally, IP-10 stimulated the ex vivo recruitment of CXCR3+ effector T cells as well as CXCR3+ Tregs derived from patients with PDAC. Our findings suggest that, in pancreatic cancer, CXCR3+ Tregs can be recruited by IP-10 expressed by PSCs in the tumor stroma, leading to immunosuppressive and tumor-promoting effects.

Optimal duration and timing of adjuvant chemotherapy after definitive surgery for ductal adenocarcinoma of the pancreas: ongoing lessons from the ESPAC-3 study.

PURPOSE: Adjuvant chemotherapy improves patient survival rates after resection for pancreatic adenocarcinoma, but the optimal duration and time to initiate chemotherapy is unknown. PATIENTS AND METHODS: Patients with pancreatic ductal adenocarcinoma treated within the international, phase III, European Study Group for Pancreatic Cancer-3 (version 2) study were included if they had been randomly assigned to chemotherapy. Overall survival analysis was performed on an intention-to-treat basis, retaining patients in their randomized groups, and adjusting the overall treatment effect by known prognostic variables as well as the start time of chemotherapy. RESULTS: There were 985 patients, of whom 486 (49%) received gemcitabine and 499 (51%) received fluorouracil; 675 patients (68%) completed all six cycles of chemotherapy (full course) and 293 patients (30%) completed one to five cycles. Lymph node involvement, resection margins status, tumor differentiation, and completion of therapy were all shown by multivariable Cox regression to be independent survival factors. Overall survival favored patients who completed the full six courses of treatment versus those who did not (hazard ratio [HR], 0.516; 95% CI, 0.443 to 0.601; P < .001). Time to starting chemotherapy did not influence overall survival rates for the full study population (HR, 0.985; 95% CI, 0.956 to 1.015). Chemotherapy start time was an important survival factor only for the subgroup of patients who did not complete therapy, in favor of later treatment (P < .001). CONCLUSION: Completion of all six cycles of planned adjuvant chemotherapy rather than early initiation was an independent prognostic factor after resection for pancreatic adenocarcinoma. There seems to be no difference in outcome if chemotherapy is delayed up to 12 weeks, thus allowing adequate time for postoperative recovery.

Pancreatic cancer hENT1 expression and survival from gemcitabine in patients from the ESPAC-3 trial.

BACKGROUND: Human equilibrative nucleoside transporter 1 (hENT1) levels in pancreatic adenocarcinoma may predict survival in patients who receive adjuvant gemcitabine after resection. METHODS: Microarrays from 434 patients randomized to chemotherapy in the ESPAC-3 trial (plus controls from ESPAC-1/3) were stained with the 10D7G2 anti-hENT1 antibody. Patients were classified as having high hENT1 expression if the mean H score for their cores was above the overall median H score (48). High and low hENT1-expressing groups were compared using Kaplan-Meier curves, log-rank tests, and Cox proportional hazards models. All statistical tests were two-sided. RESULTS: Three hundred eighty patients (87.6%) and 1808 cores were suitable and included in the final analysis. Median overall survival for gemcitabine-treated patients (n = 176) was 23.4 (95% confidence interval [CI] = 18.3 to 26.0) months vs 23.5 (95% CI = 19.8 to 27.3) months for 176 patients treated with 5-fluorouracil/folinic acid (χ(2) 1=0.24; P = .62). Median survival for patients treated with gemcitabine was 17.1 (95% CI = 14.3 to 23.8) months for those with low hENT1 expression vs 26.2 (95% CI = 21.2 to 31.4) months for those with high hENT1 expression (χ(2)1= 9.87; P = .002). For the 5-fluorouracil group, median survival was 25.6 (95% CI = 20.1 to 27.9) and 21.9 (95% CI = 16.0 to 28.3) months for those with low and high hENT1 expression, respectively (χ(2)1 = 0.83; P = .36). hENT1 levels were not predictive of survival for the 28 patients of the observation group (χ(2)1 = 0.37; P = .54). Multivariable analysis confirmed hENT1 expression as a predictive marker in gemcitabine-treated (Wald χ(2) = 9.16; P = .003) but not 5-fluorouracil-treated (Wald χ(2) = 1.22; P = .27) patients. CONCLUSIONS: Subject to prospective validation, gemcitabine should not be used for patients with low tumor hENT1 expression.

Effect of adjuvant chemotherapy with fluorouracil plus folinic acid or gemcitabine vs observation on survival in patients with resected periampullary adenocarcinoma: the ESPAC-3 periampullary cancer randomized trial.

CONTEXT: Patients with periampullary adenocarcinomas undergo the same resectional surgery as that of patients with pancreatic ductal adenocarcinoma. Although adjuvant chemotherapy has been shown to have a survival benefit for pancreatic cancer, there have been no randomized trials for periampullary adenocarcinomas. OBJECTIVE: To determine whether adjuvant chemotherapy (fluorouracil or gemcitabine) provides improved overall survival following resection. DESIGN, SETTING, AND PATIENTS: The European Study Group for Pancreatic Cancer (ESPAC)-3 periampullary trial, an open-label, phase 3, randomized controlled trial (July 2000-May 2008) in 100 centers in Europe, Australia, Japan, and Canada. Of the 428 patients included in the primary analysis, 297 had ampullary, 96 had bile duct, and 35 had other cancers. INTERVENTIONS: One hundred forty-four patients were assigned to the observation group, 143 patients to receive 20 mg/m2 of folinic acid via intravenous bolus injection followed by 425 mg/m2 of fluorouracil via intravenous bolus injection administered 1 to 5 days every 28 days, and 141 patients to receive 1000 mg/m2 of intravenous infusion of gemcitabine once a week for 3 of every 4 weeks for 6 months. MAIN OUTCOME MEASURES: The primary outcome measure was overall survival with chemotherapy vs no chemotherapy; secondary measures were chemotherapy type, toxic effects, progression-free survival, and quality of life. RESULTS: Eighty-eight patients (61%) in the observation group, 83 (58%) in the fluorouracil plus folinic acid group, and 73 (52%) in the gemcitabine group died. In the observation group, the median survival was 35.2 months (95%% CI, 27.2-43.0 months) and was 43.1 (95%, CI, 34.0-56.0) in the 2 chemotherapy groups (hazard ratio, 0.86; (95% CI, 0.66-1.11; χ2 = 1.33; P = .25). After adjusting for independent prognostic variables of age, bile duct cancer, poor tumor differentiation, and positive lymph nodes and after conducting multiple regression analysis, the hazard ratio for chemotherapy compared with observation was 0.75 (95% CI, 0.57-0.98; Wald χ2 = 4.53, P = .03). CONCLUSIONS: Among patients with resected periampullary adenocarcinoma, adjuvant chemotherapy, compared with observation, was not associated with a significant survival benefit in the primary analysis; however, multivariable analysis adjusting for prognostic variables demonstrated a statistically significant survival benefit associated with adjuvant chemotherapy. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00058201.