In the Neoarchean (~ 2.7 Ga) contact metamorphosed charnockitic footwall of the Mesoproterosoic (1.1 Ga) South Kawishiwi intrusion of the Duluth Complex, the primary metamorphic mineral assemblage and Cu–Ni–PGE sulfide mineralization is overprinted by an actinolite + chlorite + cummingtonite + prehnite + pumpellyite + quartz + calcite hydrothermal mineral assemblage along 2–3 cm thick veins. In calcite, hosted by the hydrothermal alteration zones and in a single recrystallized quartz porphyroblast, four different fluid inclusion assemblages are documented; the composition of these fluid inclusions provide p–T conditions of the fluid flow, and helps to define the origin of the fluids and evaluate their role in the remobilization and reprecipitation of the primary metamorphic sulfide assemblage.2 fluid inclusions were found as early inclusions in recrystallized quartz porphyroblast. These inclusions may have been trapped during the recrystallization of the quartz during the contact metamorphism of the footwall charnockite in the footwall of the SKI. The estimated trapping pressure (1.6–2.0 kbar) and temperature (810–920 °C) conditions correspond to estimates based on felsic veins in the basal zones of the South Kawishiwi intrusion.Pure CO2–H2O–NaCl and CH4–N2–H2O–NaCl compositions found in this study along healed microfractures in the recrystallized quartz porphyroblast establish the heterogeneous state of the fluids during entrapment. The estimated trapping pressure and temperature conditions (240–650 bar and 120–150 °C for CO2–H2O–NaCl inclusions and 315–360 bar and 145–165 °C for CH4–N2–H2O–NaCl inclusions) are significantly lower than the p–T conditions (> 700 °C and 1.6–2 kbar) during the contact metamorphism, indicating that this fluid flow might not be related to the cooling of the Duluth Complex and its contact aureole. The presence of chalcopyrite inclusions in these fluid inclusions and in the trails of these fluid inclusion assemblages confirms that at least on local scale these fluids played a role in base metal remobilization. No evidences have been observed for PGE remobilization and transport in the samples. The source of the carbonic phase in the carbonic assemblages  could be the graphite, present in the metasedimentary hornfelsed inclusions in the basal zones of the South Kawishiwi intrusion.Fluid inclusion assemblages with CO2 equiv. wt.%), low temperature (90–160 °C), primary aqueous inclusions were found. Chalcopyrite , replacing and intersecting the early hydrothermal phases, are associated to the late calcite (II) phase. The composition of the formational fluids in the Canadian Shield is comparable with the composition of the studied fluid inclusions. This suggests that the composition of the fluids did not change in the past 2 Ga and base metal remobilization by formational fluids could have taken place any time after the formation of the South Kawishiwi intrusion.The hydrothermal veins in the charnockite can be characterized by an actinolite + cummingtonite + chlorite + prehnite + pumpellyite + calcite (I–II) + quartz mineral assemblage. Chlorite thermometry yields temperatures around 276–308 °C during the earliest phase of the fluid flow. In the late calcite (II) phase, high salinity (21.6–28.8 NaCl + CaCl34S = 7.4–8.9‰) and the hydrothermal sulfide mineral assemblage (δ34S = 5.5–5.7‰) proves, that during the hydrothermal fluid flow the primary metamorphic ores were remobilized.Sulfur isotope studies carried out on the primary metamorphic  transported copper.Mixing carbonic–aqueous fluids (CO•High-salinity calcium-rich basinal brines remobilized copper and nickel. The volumetrically important base and precious metal ores in these intrusions were produced by segregation, fractionation and settling of immiscible magmatic sulfide , but othIt has been presented recently that high grade, though small volume Cu–Ni and PGE mineralization may also occur in the footwall lithologies of layered intrusions  of the South Kawishiwi intrusion (SKI) by 18O for H2O) they proved that fluids both from magmatic and metasedimentary footwall sources were involved in the Pt–Pd redistribution. These studies have the following economic significance: 1.) fluid circulation in the primary mineralized zones of the intrusion may have resulted in depletion of the ores in PGEs, and 2.) there is a high probability of the formation of secondary hydrothermal Cu-mineralization also far from the basal mineralized zones, in the hanging wall or in the footwall units of the intrusion.Hydrothermal processes and their roles in PGE remobilization in the troctolitic intrusions of the DC have been demonstrated by The aim of this paper is to present the textural, mineralogical and geochemical characteristics of the hydrothermal alteration zones in the charnockitic footwall of the SKI and to characterize the hydrothermal fluids based on fluid inclusion studies. Fluid inclusion studies have not only been carried out on footwall samples, but also on non-metamorphosed and unaltered granitoid samples far from the contact aureole, in order to distinguish regional and local fluid flow events.22.12 arcuate area in northeastern Minnesota (The DC and associated intrusions in northeastern Minnesota are part of the Mesoproterozoic (1.1 Ga) Midcontinent Rift MCR; A which iinnesota B. The DCinnesota .The DC is in its current position tilted by 15–20° to SE, and therefore the immediate footwall units of the SKI, Bathtub intrusion and Partridge River intrusion crop out along the north-eastern perimeter of the DC.2.2Footwall rocks adjacent to and beneath the DC include Neoarchean intrusive, sedimentary and magmatic rocks and Paleoproterozoic sedimentary rocks of the Animikie Basin. Neoarchean granitoid rocks form the direct footwall with mineralized layered intrusions only the northeastern segment of the SKI. Granitic, granodioritic, monzonitic and tonalitic rocks comprise most of the 2.7 Ga Giants Range batholith (GRB), which is intrusive into the supracrustal rocks of the Wawa–Abitibi subprovince — the southernmost granite–greenstone belt of the Superior Province .Except for a short (~ 25 km in strike length) segment along the SKI, the footwall of the DC is the Early Proterozoic Biwabik Iron Formation and the graphite- and pyrite-bearing Virginia Formation. Multiple intrusions of cumulate textured anorthositic and troctolitic rocks produced high-temperature amphibole–pyroxene hornfels facies contact metamorphism, devolatilization and locally partial melting of the footwall lithologies .2.3The dominantly disseminated Cu–Ni–PGE sulfide mineralization of the SKI, Partridge River intrusion and Bathtub intrusions C is conf34S values range from 0 to 26‰ and the sulfur in the basal zones of the Partridge River intrusion and the Bathtub intrusion is derived from the metasedimentary footwall. 2 or H2S. In addition — due to the fluid–rock interaction — the organic component (graphite) in the metasedimentary Virginia Formation liberated in the form of CO2 and CH4 during the contact metamorphism by the DC  resulted in intense mineralogical, textural and geochemical modifications of the footwall. According to the studies of y the DC . The SO2al melts  migratedal melts . Upward The late-stage, low-temperature (< 350 °C) hydrothermal fluid circulation resulted in local formation of actinolite + prehnite + pumpellyite + chlorite + calcite veins and alteration zones in the SKI and in the Partridge River intrusion . These f2 ± CH4 ± N2 and NaCl + CaCl2 + KCl) and with highly variable salinities (from 0 to 48 wt.% NaCl + CaCl2) and emphasized the potential of brine–gas and brine–brine immiscibility in modification of primary distribution of metals. 2 (± CH4 ± N2) inclusions in local felsic segregations and cross-cutting felsic dikes presumably originated from partial melting of footwall in the SKI and suggested that the CO2-rich deuteric fluid phase originated from the mafic magma.There are only a few studies dealing with characterization of hydrothermal fluids affecting DC mineralized zones by fluid inclusion studies. 2.4Granitoid rocks of the GRB beneath the SKI belong to the Farm Lake Facies, which is a porphyritic hornblende- or biotite-bearing monzodiorite that consists of plagioclase + K-feldspar + quartz + biotite + hornblende ; Fig. 1DAccording to the recent studies of No metal concentration data are available from the studied WM-002 drill core, but both the intrusion and the footwall part of the near-by WM-001 drill core was systematically analyzed . Metal c3Samples were collected from the drill core library of the Minnesota Department of Natural Resources in Hibbing. Two drill cores the WM-001 and WM-002, acquired by Wallbridge Mining from the Spruce Road area  C and D Electron microprobe analysis on polished thin sections was performed on a JEOL 6310 SEM, equipped with a Link ISIS energy dispersive system and a Micro-Spec wavelength dispersive system at the University of Graz. Accelerating voltage was 15 kV, the probe current 5 nA for silicates and oxides. Detection limits are 0.1–0.2 wt.% for EDS and 0.03 wt.% for WDS analysis.2O5 was added to the samples as a combustion aid. δ34S results are reported in units of ‰ vs. VCDT. An uncertainty of ± 0.2‰ is recommended.Sulfur isotope analyses were carried out in the isotope lab of the Department of Geological Sciences and Engineering, University of Nevada, Reno. Analyses were performed using a Eurovector EA 3000 elemental analyzer interfaced to a Micromass IsoPrime stable isotope ratio mass spectrometer, after the methods of Fluid inclusions have been investigated in oriented doubly-polished thick sections (thickness ca. 0.15 mm). Measurements of phase transition temperatures in fluid inclusions were performed using a LINKAM THSMG600 heating and freezing stage with an operating range from − 196 °C to + 600 °C , equippe2, CH4 and N2 bearing inclusions from microthermometric data  the module BULK of the Software Package Fluids v.1 was used  chlorite replaces rock-forming metamorphosed oligoclase of the parent charnockite host and actinolite that formed from the alteration assemblage affecting metamorphic pyroxene E. That cWhere the fluids of the second hydrothermal assemblage affected the sulfide assemblage that filtrated during the contact metamorphism in the partially molten charnockite, the pyrrhotite and the chalcopyrite are intensively corroded and occur as inclusions in the pyrite E. Pyrite4.1.2In the WM-002 drill core, beside the calcite crystals in the hydrothermal veins, only one relict quartz porphyroblast, 10 m beneath the intrusion–footwall contact was found suitable for fluid inclusion studies .The porphyroblast consists of a polygonal aggregate of quartz crystals, without internal stress features . The crystals have straight or slightly rounded grain boundaries and a 120° triple junction between grains, indicating equilibrium recrystallization.The mechanism of formation of the porphyroblast is enigmatic, since the modal proportion of the quartz in the host partially molten charnockite is low (< 1%). Quartz occurs only as cuspate formed crystals between feldspar porphyroblasts  or in hiThe porphyroblast is mantled by a 1 mm thick halo composed of clinopyroxene, orthopyroxene, and chalcopyrite + pyrrhotite with minor galena and sphalerite as accessories A. The quHowever, only one single quartz porphyroblast was found in the drill core, this single crystal can be representative since it is mantled by ortho- and clinopyroxene that formed during the peak metamorphism. Therefore, the quartz has probably also recrystallized during the metamorphism. On the other hand this is the only mineral in the drill cores that may have preserved fluids that were present during the peak of the contact metamorphism.4.2apfu)] and total Fe [= Fe2 + + Fe3 + (apfu)] values ranging from 2.53 to 4.84 (apfu). The Fe/(Fe + Mg) ratio varies from 0.26 to 0.5; the Fe3 + value is very low, characterizing the chlorites to be Fe-rich and unoxidized. The Si/Al ratio shows very little variability, ranging from 1.18 to 1.27 and the Al in the tetrahedral site varies between 2.53 and 2.84 ions per formula unit  represent the samples from the footwall with actinolite–chlorite–quartz–calcite–prehnite–pumpellyite alteration.In order to compare isotopic variations during magmatic, metamorphic and hydrothermal processes, the following mineralization types were sampled in the drill cores in this study: (1) disseminated type chalcopyrite–cubanite–pentlandite ore in the basal mineralized zone, (2) semi-massive pyrrhotite–pentlandite–chalcopyrite dominated mineralized basal zone of the intrusion, (3) chalcopyrite-rich chlorite–calcite–prehnite veins and patches in the troctolite, (4) pyrrhotite–pentlandite–chalcopyrite dominated sulfide droplets that percolated into the proximal zones of the footwall during peak metamorphism, (5) pyroxenite-veins with chalcopyrite and pentlandite, (6) bornite–chalcopyrite composite grains associated with quartz–biotite melt patches and (7) chalcopyrite-rich ore associated to chlorite–quartz–calcite–prehnite–pumpellyite alteration assemblage. Results of sulfur isotope analyses are listed in 4.44.4.1Type ICarbonic inclusions were observed in the quartz–porphyroblast, 10 m below the intrusion–footwall contact. The fluid inclusions occur isolated, or randomly in three-dimension arranged clusters A. These Type II2–H2O–NaCl system with highly variable phase ratios are arranged along fluid inclusion planes (FIPs) in the recrystallized quartz porphyroblast. At room temperature, the inclusions have three phases, with highly variable volume fractions; carbonic vapor (Vcar), carbonic-liquid (Lc) and aqueous-liquid (Laq) (These types of inclusions belonging to the COid (Laq) B–C. Optiid (Laq) C–D.Type III4–N2–H2O type composition, and these inclusions occur along fluid inclusion planes in the recrystallized quartz porphyroblast and in apatite, which host the actinolitic alteration zones. They define one phase vapor or two phase liquid + vapor inclusions  as primary and secondary inclusions in calcite II that is intergrown with prehnite and pumpellyite in the hydrothermal veins  asal veins C; and (3Four different fluid inclusion assemblages (FIA) have been distinguished on the basis of the host mineral, petrographic location and the microthermometric behavior. Petrographic and microthermometric properties of the different FIA and the calculated salinities are summarized in 4.4.2Results of fluid inclusion microthermometry and the calculated salinities are summarized in Type I FIA melting temperatures of CO2 is close to − 56.6 °C (− 56.7 °C to − 56.6 °C) indicating pure CO2 chemistry. The inclusions homogenize into the liquid phase between 30.7 °C and 31.0 °C (3/mol) and the density range (0.51–0.55 g/cm3) for the inclusions were calculated from the homogenization temperatures.The  31.0 °C A. The buType II FIA melting temperatures of the CO2 in the Lcar-rich inclusions is − 56.6 °C, indicating no other volatiles (CH4 or N2) than CO2 are in the inclusions. Observation of eutectic melting around ~ 21 °C was possible only in one inclusion due to the small size of the aqueous inclusions or due to the low H2O/CO2 ratio. Ice melting and clathrate melting temperatures varied from − 3.9 °C to − 3.2 °C and from 8.4 to 8.9 °C, respectively. Salinities calculated from the clathrate melting temperatures vary from 1.8 NaCl equiv. wt.% to 2.7 NaCl equiv. wt.%. Homogenization temperatures of the carbonic phase into Lcar range from 24.7 °C to 31 °C , and the homogenization temperature of the Lm and Vm into Vm phase varies between − 121 °C and − 101.4 °C (4 (− 82.6 °C) and pure N2 (− 147.0 °C) system. According to the analysis of the Raman spectra, using the method of 4/N2 ratio shows large variation between 0.15 and 1 with an average of 0.3. Molar volume and density for the gas-rich end member vary from 112.9 cm3/mol to 119.6 cm3/mol and from 0.21 g/cm3 to 0.23 g/cm3, respectively. Melting temperatures of ice phase in the aqueous-rich end members vary between − 0.7 °C and − 0.1 °C. Salinities vary between 0.23 and 1.25 NaCl equiv. wt.%, assuming an average CH4/N2 ratio of 0.15 and an average homogenization temperature of − 103 °C. Bulk molar volumes for aqueous end members vary from 19.51 mol/cm3 to 19.55 mol/cm3 and the densities range from 0.92 g/cm3 to 0.931 g/cm3. Homogenization temperatures of the aqueous end members vary between 126 °C and 251 °C (101.4 °C B. These d 251 °C C and theType IV FIA eutectic temperatures (from − 52.3 °C to − 48.6 °C) were detected close to the eutectic temperature (~− 52 °C) for the H2O–NaCl–CaCl2 system. Melting of the hydrohalite and ice phases were observed from − 36.3 °C to − 19.4 °C and from − 27.8 °C to − 12.3 °C, respectively. Salinities range from 21.6 to 28.8 NaCl + CaCl2 equiv. wt. % are calculated from the hydrohalite and ice melting temperatures (2/(CaCl2 + NaCl) ratio range between 0.3 and 0.6. Homogenization of the inclusions occurred with the disappearance of the vapor phase (L + V → L) between 84.5 and 166.2 °C. In the uncontaminated rock forming quartz of the footwall granite and prehnite–pumpellyite–quartz–calcite alteration zones in the WM-002 drill core, homogenization temperatures range from 160 °C to 170 °C and between 110 °C and 120 °C, respectively .Peak metamorphic temperatures calculated for contact metamorphic conditions, based on two-pyroxene thermometry  the overburden during the peak metamorphism could be 5.4–6.7 km. This thickness is much higher than the current location of the sample below the surface (300 m), but less than the estimated total thickness of the DC  the thickness of the overburden rock thickness can be calculated. Assuming lithostatic conditions (3000 kg/m5.1.24–N2 inclusions), the coexistence of gas-rich and aqueous liquid-rich inclusions with various degree of filling indicate either: (1) heterogeneous entrapment in an immiscible fluid system, or (2) mixing of two fluids with different compositions (2–CH4–N2–H2O). Therefore, mixing or partial mixing of the Type II and III FIA can be excluded. The two FIA, therefore, indicate two different fluid mixing events.In the fluid inclusion planes of the Type II FIA (carbonic–aqueous inclusions) and Type III  and the carbonic-rich (Thcar = 24 to 31 °C) end members . The pre4–N2–H2O) inclusions also displays heterogeneous trapping. Similar to the Type II assemblage, mixing of two primarily homogeneous fluid phases is assumed. The representative isochors of gas-rich inclusions are calculated for molar volumes of 113 and 120 mol/cm3, and those of the aqueous-rich inclusions for Th = 130 °C and 150 °C. The isochors' intercept is in the range between 315 and 360 bar and 145 °C to 165 °C .2O (deuterium-oxide) and re-equilibrate. Hydrogen diffusion, as it was suggested by 2 inclusions or precipitation of graphite and hydrogen in methane-bearing inclusions. Loss of water from carbonic inclusions may also trigger precipitation of graphite. Neither graphite or hydrogen in the inclusions or inclusions with variable CO2/CH4 ratios were detected, therefore, hydrogen diffusion or selective water leakage likely did not change the bulk composition of the inclusions.To explain these discrepancy in the pressure and temperature conditions, three scenarios are proposed: (1) selective diffusion of volatiles through the fluid inclusion wall after entrapment that changes the primary bulk composition , or (2) diagenetic origin of fluids. The first process has been proposed by According to the second scenario, the metasedimentary Virginia Formation or the hornfelsed Virginia Formation inclusions in the SKI may have released carbonic-rich fluids due to the groundwater–rock interaction any time after the formation of the SKI. These fluids may have migrated in the SKI and in the neighboring rock volumes and along some preferred permeable zones. This latter model also explains why no volatile-rich fluids were encountered far (~ 10 km) from the contact in the GRB.5.2The actinolite + cummingtonite + chlorite + albite + quartz + calcite + prehnite + pumpellyite  assemblage corresponds to the characteristic mineral assemblages of lower greenschist–prehnite–pumpellyite facies  in the crystal structure was first reported by IV vs. Fe/(Fe + Mg) and the Si/Al vs. Fe/(Fe + Mg) ratios (both given in apfu) show that the analyzed samples are comparable with the composition fields of A systematic relationship between the formation temperature of chlorite and the Al2 is lower than 0.01 in the system. Even if the isochors of these inclusions are intercepted with the 1.7 kbar isobar (trapping pressure of Type I primary fluid inclusions), the trapping temperatures do not exceed the 200 °C. Therefore, calcite II can be considered as the last phase to precipitate during hydrothermal fluid flow.The Type IV FIA does not show any evidence of heterogeneous trapping. Therefore the homogenization temperatures (110–120 °C and 160–170 °C) represent the minimum temperature of trapping. In order to obtain the trapping pressures, an independent thermometer, or barometer is required. A possible candidate would be the prehnite with which the calcite II is apparently syngenetic, but prehnite precipitates only if the activity of the CO5.35.3.12-bearing inclusions are common in charnockites  mantle degassing, (2) decarbonation of carbonate-bearing lithologies , or (3) 2 migration in charnockitic rocks. They found that percolation of CO2 along grain boundaries is inhibited due to the large wetting angles but possible and fast (338 mm in 500 years) along microfractures. CO2 diffusion in melts, however, is only possible above ~ 840 °C. Considering that the temperature in the proximal 10 m part of the footwall of the DC was between 830 and 910 °C and the granite was in a partially molten state, both processes could play role in the CO2 diffusion into the granite, but possibly the presence of high permeable zones (pyroxene-rich veins) promoted the CO2 diffusion.Recently, 5.3.22 wt.%) of regional formational fluids do not show any significant variation in the past 2 Ga years. The reported compositions are comparable with the compositions of the fluid inclusions discussed in this work.Calcium-bearing, high salinity fluids (Type IV) have been found as primary inclusions in calcite II in the hydrothermal vein assemblage and in the rock forming quartz of the GRB 10 km away from the intrusion–footwall contact. Ca-rich high salinity fluids are commonly found in sedimentary basins and on shields where the high salinity is the result of the significant fluid–rock interaction under low fluid/rock ratio conditions. Therefore, these fluids are considered as regional formational fluids. Groundwater in the Soudan Mine (~ 30 km away from the Duluth Complex) has been recently investigated by 5.434S) of sulfide assemblages related to the partial melting have average values of approximately + 8‰ in the footwall, and are comparable with the sulfide isotope ratios in the basal zones of the intrusion (this study). The similar values indicate that during the infiltration of the sulfide liquid into the footwall, along some permeable zones created by the partial melting  isotopic fractionation is minor and hence can be neglected e.g., . Howeverhe fluid . Accordial fluid . We prop34S of HS− and H2S (aq) to lighter values vs. total dissolved sulfur, and hence δ34S value of sulfide minerals in the hydrothermal veins will decrease vs. sulfur in the hydrothermal fluid  and CH4 (Type III) bearing fluids only in the recrystallized quartz porphyroblast, which is mantled by chalcopyrite and pyrrhotite. Therefore, the estimation of the extent of this remobilization is difficult.Chalcopyrite inclusions in the Type II and III fluid inclusion assemblages indicate that these fluids played a role in metal redistribution. However, these solid inclusions are found associated with COHydrothermal veins with comparable mineral assemblages have been reported from the Layered and Anorthositic Series of the Partridge River intrusion and SKI . These sHydrothermal alteration assemblages and Ca-bearing fluids have been found in the SKI, Partridge River intrusion and Bathtub intrusion  of the D62-bearing fluid inclusions were found as primary inclusions in a recrystallized quartz porphyroblast with near critical homogenization temperatures (30.7–31 °C). Assuming that these inclusions trapped during the peak metamorphism when the temperatures were in the range from 830 to 920 °C, the calculated pressure of trapping varies between 1.6 and 2.0 kbar. This pressure range corresponds to earlier pressure estimations, carried out in the felsic dykes at the bottom of the SKI  to low temperatures (< 200 °C). Composition of fluid inclusions  found in the second generation of calcite are comparable with current groundwater compositions and hence involvement of shield brines in the late stage fluid–rock interaction related to the cooling of the contact aureole of the Duluth Complex. Since Ca-bearing high-salinity fluids resulted in alterations with similar alteration assemblages in several units of the Duluth Complex, the possibility of a post-Keweenawan fluid flow event cannot be excluded.34S ≈ 8‰) infiltrated primarily during the contact metamorphic process from the intrusion into the footwall. During the hydrothermal remobilization due to the preferred fractionation of 32S in the sulfide phase, the δ34S values will decrease compared to the source sulfur isotope values. Accordingly, the measured low δ34S values range around + 5.5‰.Sulfur isotope studies have revealed that the different sulfide assemblages characterized by sulfur isotope values (δ


The amount of fluid during resuscitation of septic shock is important. Too little fluid may result in tissue hypoperfusion; however, too much fluid may result in volume accumulation. Several recent studies have demonstrated that a positive fluid balance in critical illness is associated with deteriorating outcomes. However, some studies have shown opposite results. The objective of this study was to determine whether initial fluid balance in septic shock patients is correlated with ICU mortality.This is a retrospective study of septic shock patients admitted to a mixed medical-coronary care unit of Songklanagarind hospital from 2005 to 2011. Multivariate logistic regression analysis was used to identify predictors of mortality.n = 555 (53%)) and nonsurvivors (n = 493 (47%)). Median survival time was 10 days (95% CI: 8 to 12 days). The respiratory tract was the most common site of infection (47.6%). Community-acquired infections accounted for 59.6%. Survivors were older than nonsurvivors . Nonsurvivors were more severely ill and had shorter ICU stays . Nonsurvivors received albumin and steroid more than survivors. Median cumulative fluid at 24, 48 and 72 hours of septic shock onset were 4.2, 7.7 and 10.5 l respectively. Nonsurvivors had significantly larger median cumulative fluid intake at 24 hours , at 48 hours  and at 72 hours . Nonsurvivors also had significantly larger fluid balance  and mean fluid balance  within 72 hours. In multivariate logistic regression analysis, factors significantly associated with ICU mortality were mean fluid balance, APACHE II score, SOFA score, length of ICU stay, ARDS, steroid use, parenteral nutrition use and source of infection.A total of 1,048 patients admitted to ICU for septic shock was divided into two groups: in-ICU survivors (A more positive cumulatively fluid balance over 3 days is associated with ICU mortality in septic shock. Multivariate analysis found not only nonmodifiable factors such as severity score, source of infection, length of ICU stay and ARDS, but also modifiable factors such as parenteral nutrition use, steroid use and mean fluid balance were significantly associated with mortality.


Colorectal cancer is the third most common cause of cancer-related deaths and the second most prevalent (after breast cancer) in the western world. High metastatic relapse rates and severe side effects associated with the adjuvant treatment have urged oncologists and clinicians to find a novel, less toxic therapeutic strategy. Considering the limited success of the past clinical trials involving peptide vaccine therapy to treat colorectal cancer, it is necessary to revise our knowledge of the immune system and its potential use in tackling cancer. This review presents the efforts of the scientific community in the development of peptide vaccine therapy for colorectal cancer. We review recent clinical trials and the strategies for immunologic monitoring of responses to peptide vaccine therapy. We also discuss the mechanisms underlying the therapy and potential molecular targets in colon cancer. Colorectal cancer (CRC) is the third most common cause of cancer-related deaths among men and women in the western world ,2. EuropSince the discovery of tumour antigens in 1991, immunotherapy has become a potential alternative to conventional chemotherapy . In 2010Immunotherapy is based on the principle of aberrant expression of proteins by tumours, which either over-express ubiquitous proteins or express proteins that unusually occur in the original tissue of the cancer. Tumour cells that bear these antigens can be distinguished from the normal tissue by the immune system in the same way that bacteria or virus-infected cells are. Recognition of an unusual antigen starts an immune response aimed at elimination of the cell and generation of antigen-specific immune cells that provide long-lasting immunity. However, immune-mediated regression of cancer hardly ever takes place in real life due to the low effectiveness of wild-type antigens (generated by the tumour) in stimulating host immune responses. The aim of vaccine therapy is to provide a highly immunogenic antigen, such as peptides, which are capable of stimulating immune system to mount a cytotoxic attack against the tumour cells. The desired effect would be stabilization or even regression of the disease.Following an inoculation, the circulating peptides are degraded in the circulation and the antigenic products are endocytosed by antigen-presenting cells (APC), which migrate to lymph nodes where they present the antigen in a processed form to T cells. These are at the forefront of the battle against the tumour. There are two main types of T cells according to the surface markers and function. CD8+ T cells, also known as cytotoxic T lymphocytes (CTL), are responsible for direct killing of tumour cells. They induce apoptosis by release of granzymes and perforins or by presenting Fas ligand (FasL) to Fas death receptor on the target cells. Another consequence of granzyme release is target cell necrosis [The interaction between individual cells is mediated by two major histocompatibility complex (MHC) proteins, MHC classes I and II , present on the surface of cells. The antigenic protein from the circulation is processed by cellular machinery into a several-amino acid-peptide that can enter the MHC molecule for presentation to T cell receptor (TCR). ●MHC class I is responsible for presentation of the vaccine-derived peptide between APC and naïve CTLs. Primed CTLs are able to recognize the genuine tumour antigen presented by the MHC I on the surface of the tumour. Thus activated CTL sends out a death signal to the tumour.●MHC class II is responsible for “the talk” between APC and CD4+ T cells and subsequent generation of helper T cells. Cytokines secreted by helper T cells, such as IL-2, are essential for activation of strong cytotoxic pathways and potentiation of anti-tumour response [response . All in Various immune cells are normally found infiltrating the mass of a tumour, some of which are specialized to recognize the tumour-associated antigens. Although there is evidence of a positive correlation between the number of these cells and good prognosis, their effect on the tumour is considered insignificant ,11. It h●One such mechanism is expression of FasL, which is a ligand for Fas receptor present on CTL. Fas functions as a death receptor which, upon binding of its ligand expressed on the tumour surface, induces CTL to undergo apoptosis. Additionally, expression of FasL provides a degree of resistance to Fas-induced apoptosis in tumour cells themselves .●Another mechanism involves expression of altered peptide ligands for T cell receptors (TCR) by the tumour. “Altered Peptide Ligand” (APL) are analogs of immunogenic peptides in which the TCR contact sites have been manipulated . Recogni●Oncogenic signalling pathways within tumour cells and immunologic checkpoints in the tumour microenvironment also play a crucial role in promoting immunologic tolerance. For example, tumour releases factors that induce inhibition of both innate and adaptive antitumor immunity. Stat3 activation in tumours, as well as Braf activation, can induce release of factors such as IL10 that induce Stat3 signalling in NK cells, granulocytes, inhibiting their tumouricidal activity. Stat3 is also activated within conventional dendritic cells (CDC) in the tumour, converting them to toleragenic DC, which can induce T cell anergy and possibly regulatory T cells (Treg). Plasmacytoid DC (PDC) or PDC-related cells in the tumour microenvironment upregulate indoleamine 2,3-dioxygenase (IDO), an enzyme that metabolizes tryptophan. T cells are very sensitive to tryptophan depletion. Tumours can express co-inhibitory B7 family members, such as B7-H1 and B7-H4, which downregulate T cell activation and/or cytolytic activity. They can also induce B7-H1 and B7-H4 expression on tumour associated macrophages (TAM). Related immature myeloid cells or myeloid suppressor cells can further inhibit antitumor T cells via production of NO by the enzyme arginase. ,17.●Regulatory T cells are an important inhibitor of antitumor immunity. T cell activation in the absence of appropriate co-stimulatory signals leads to T cell anergy and generation of induced regulatory T cells (Treg). Treg, characterized by the FoxP3 transcription factor, upregulate a number of cell membrane molecules, including Lag3, CTLA4, GITR, and neuropilin. Treg can inhibit effector T cell activation and function via T-T inhibition or inhibition of antigen presenting cells .●Finally, cancers avoid recognition by the immune system by means of defective antigen presentation. This can be achieved by reduced expression of HLA type I (MHC I), which is a common event in colorectal cancer , as wellImmune-competence of cancer patients who receive chemotherapy is already partially compromised. The design of peptide vaccines is aimed at overcoming the described problems by delivering highly immunogenic antigens combined with adjuvants that stimulate the immune system.Special design of the amino acid sequence of a peptide can enhance the interaction with the TCR. This usually involves creation of a peptide epitope with single amino acid substitutions, which exhibits improved binding to MHC or affinity for the TCR. Such manipulations are capable of potentiating secretion of interleukin 2 , which is a potent immune-boosting cytokine. Other modifications, involving substitution at the peptide terminals, results in improved bio-stability and reduced degradation by seric proteases . EffectiThe use of synthetic peptides is a fairly recent endeavour. In the early days of immune therapy tumour antigens were delivered to the patient in a form of tumour lysates. Nowadays tumour lysates are being pulsed into dendritic cells (DC) as a way of facilitating peptide presentation. Another recent development is the use of a retrovirus carrying the antigen-encoding gene, which inserts itself in the host genome leading to production of large quantities of the antigen . AgainstThe main drawback is the human leukocyte antigen (HLA)-type restricted nature of the therapy. HLA molecules are expressed in a multitude of super-types and individual antigens are only capable of interacting with a particular HLA-super type. In the clinical context this means that only patients carrying the required HLA allele will be able to respond to the therapy. This substantially limits the application of the therapy. Moreover, use of self-antigens carries a risk of inducing autoimmunity. This kind of therapy is also contraindicated in patients with existing autoimmune diseases, where exposure of the over-reactive immune system to a highly immunogenic antigen can have unpredictable consequences. An example of that comes from trials on immune therapy in Parkinson’s disease where the patients’ condition was reported to be aggravated following vaccination against a neuroprotein .Two aspects of response to peptide therapy are commonly evaluated in clinical setting: clinical outcome as the primary endpoint and stimulation of cell-mediated toxicity as a secondary endpoint. Several techniques have been used to measure the frequency and activity of antigen-specific CTLs, including ELISPOT assay, flow cytometry, RT-PCR, HLA/epitope tetramer assay. This is often combined with evaluation by means of serum cytokines and chromium (υ¹Cr) release assays, and proliferation studies . The υ¹C●ELISPOT assays are used to assess secretion of proteins by CTLs which are indicative of an activated state, such as, INF-γ, granzyme B, IL-2 . The ass●15, EL4 and T2 lymphoma cells with a cell tracker dye DDAO-SE and staining permeabilized cells with antibody against cleaved (activated) caspase-3. This assay proved to be robust and reliable in evaluating antigen-specific CTL [Following T-cell receptor recognition of antigenic peptide–MHC class I complexes on the surface of target cells, CTLs induce target cell apoptosis through directed exocytosis of perforin and granzymes. The cytotoxic signalling leads to the activation of the caspase cascade that can be measured using flow cytometer. The assay involved labelling of P8ific CTL .●Another approach is to use flow cytometry for detection of apoptosis in CTL target cells . During apoptosis, phosphatidylserine (PS) is externalized on the surface of the cells and is available for binding of annexin V. Both granule marker and annexin V assays allow evaluation of the cytotoxic potential of tumour-specific CTLs .●By contrast, HLA/epitope tetramer assay is used for mere enumeration of the antigen-specific CTLs. However, when combined with intracellular cytokine staining it gives a complete picture of quantity and function of the TCLs.●Lastly, RT-PCR has been used to evaluate cytokine gene expression, including INF-gamma and granzyme B, by CTLs ,26. CellIdeally the immune response should correlate with the primary endpoints, which reflect the disease progression. Sadly this has been exceptionally rare in the clinical trials so far. In addition to patient survival, the levels of tumour markers are measured using fluorescence-based immunoassays for monitoring of tumour growth and response to therapy . Also, dTumour self-antigens are broadly categorized according to their specificity for the tumour. Differentiation tumour antigens present in both tumour and its original tissue, but are expressed at higher levels in the tumour. Tumour-specific antigens occur in tumours cells but are absent in normal adult tissues. Tumour/testis antigens are additionally expressed in the gonads. Unique tumour antigens constitute a separate group. These arise from nucleotide deletions or alternative splicing and characterize each single tumour. The choice of each one of these targets for vaccine therapy carries its specific risks as well as benefits. Targeting differentiation antigens, owing to their presence outside the tumour, can elicit a degree of adverse reaction in the healthy tissue. An example of that comes from clinical trials in melanoma where use of peptide vaccines directed against proteins involved in melanin biosynthesis leads to development of skin depigmentation (vitiligo). With regards to unique antigens, some evidence speaks in favour of their higher immunogenicity. The immune response against unique antigens has been found to persist for longer following tumour resection compared with shared tumour-specific antigens . Howeveri.e., melanoma. Nonetheless, the few antigen-based vaccines that have been developed show the ability to induce antigen-specific CTL and cause reduction in the tumour size in animal studies. Translation of these findings to a clinical setting was not completely successful. Tumour-specific antigens have been the most popular targets for vaccine therapy in general and in the context of colon cancer in particular. It must be noted that immunogenicity of colorectal cancers is considerably lower compared to the most immunogenic among cancers, in vivo studies and clinical trials. The remainder are summarized in the One particular pattern of response to peptide vaccine therapy prevails across all clinical studies. That is, strong induction of antigen-specific CTLs is not associated with improved clinical outcome. Frequently reported side effects of the therapy are: ulceration at the injection site, fever, fatigue, nausea, anorexia. However, serious adverse reactions are very rare. The protocols commonly require 3 or more injection courses at 2–3 weeks intervals and patients who show satisfactory response are offered continuation of the therapy. It has been argued that the lag between the first jab and the clinically relevant response is too long, especially in the case of advanced cancer patients who may not survive the necessary time. Below is a brief review of selected antigen targets in colon cancer including their biological role, results of Ephrin type-A receptor 2 (EphA2) is a member of a large tyrosine kinase receptor family. Eph receptors play an important role in oncogenesis  and tumoin vitro [Survivin is a protein which inhibits cancer cell apoptosis, is highly expressed during embryological development, and becomes undetectable in adult tissues . A studyin vitro . In 2003in vitro . The thein vitro ,48,49,50et al. used a vaccine formulation combining two SART3 antigenic epitopes recognized by HLA-A24-restricted CTLs with IFA in colorectal cancer patients. Immunization resulted in significant induction of tumour-specific CTL in 7 of 11 patients. Two patients presented with induction of INF-γ following the sixth vaccination time. However, augmentation of cellular immunity was not associated with improved clinical outcome. Likewise, no induction of IgG or IgE specific for the peptides was observed. An adverse reaction at the injection site manifested as itching and redness occurred in 6 patients. The median number of vaccination times was 8 and the median observation period was 5 months. Despite the somewhat promising result no further trials involving SART antigen has been published since 2001 [SART3 is a tumour-rejection antigen which is expressed in more than 70% colorectal cancers but is not present in normal non-malignant tissues . It was nce 2001 .Carcinoembryonic antigen (CEA) is a useful tumour marker which is correlated with tumour burden in the mouse metastasis model and has been used to monitor CRC metastasis development . A mouseOverall, immunization with CEA resulted in enhanced immune response and survival. Further improvement in both endpoints was seen with the combination therapy. This constitutes an interesting case for employment of both antibody-mediated and cell-mediated immunity to achieve greater anti-tumour response. It also consolidates the importance of T helper cells in stimulation of cytotoxic killing of the tumour cells . MUC-1 is an epithelial membrane antigen, which bound to the apical membrane of the secretory cells. It has been found that MUC-1 is expressed in more than 70% colorectal cancers and correlates with poor prognosis . An earlin vivo studies and clinical trials. The remainder are summarized in the We have discussed some of the selected antigen targets in colon cancer including their biological role, results of Development of a novel, less toxic therapeutic strategy to treat advanced and disseminated colorectal cancer has been a goal for oncologists and clinicians for many years. The discovery of tumour antigens in 1991 has opened a window for the use of immune therapy in the treatment of cancer. In the last two decades, a great deal of effort has been made to identify the molecular targets for immune therapy in colorectal cancer; some of those targets proved effective in a pre-clinical setting. Moreover, the immune therapy has been shown to cause minimal side effects in a clinical setting, which makes it a desirable alternative to the current systemic treatments. Its ability to induce strong and specific anti-tumour immune responses in patients has been well documented using the current immunologic monitoring techniques. However, these findings did not correlate with improved clinical outcome. This calls for more accurate strategies for immune monitoring and could become the next direction for research.


The accurate assessment of intravascular volume status for the therapy of severe hypovolemia and shock is difficult and critical to critically ill patients. Non-invasive evaluation of fluid responsiveness by the rapid infusion of a very limited amount of volume is an important clinical goal. This study aimed to test whether echocardiographic parameters could predict fluid responsiveness in critically ill patients following a low-volume  infusion over 10 seconds.We prospectively studied 55 mechanically ventilated patients. Echocardiography was performed during a 50-ml infusion of crystalloid solution over 10 seconds and a further 450 ml over 15 minutes. Cardiac output (CO), stroke volume (SV), aortic velocity time index (VTI), and left ventricular ejection fraction (LVEF) were recorded. Patients were classified as responders (Rs) if CO increased by at least 15% following the 500-ml volume expansion or were classified as non-responders (NRs) if CO increased by less than 15%. Area under the receiver operating characteristic curves (AUC) compared CO variations after 50 ml over 10 seconds (∆CO50) and 500 ml over 15 minutes (∆CO500) and the variation of VTI after infusion of 50 ml of fluid over 10 seconds (∆VTI50).P <0.01; best cutoff value, 6%; sensitivity, 93%; specificity, 91%). Moreover, ∆CO50 and ∆CO500 were strongly correlated . The AUC for ∆VTI50 was 0.91 ± 0.04 . ∆VTI50 and ∆CO500 were positively correlated .In total, 50 patients were enrolled, and 27 (54%) of them were Rs. General characteristics, LVEF, heart rate, and central venous pressure were similar between Rs and NRs. In the Rs group, the AUC for ∆CO50 was 0.95 ± 0.03  can accurately predict fluid responsiveness.ISRCTN10524328. Registered 12 December 2013.Current Controlled Trials  Assessment of intravascular volume status is difficult in critically ill patients. Evidence suggests that only 50% of hemodynamically unstable patients respond to a fluid challenge ,2. MoreoResearch has suggested that volume responsiveness can be defined as a 15% increase in stroke volume (SV) or cardiac output (CO) after a 500-ml infusion -12. Overet al.50 and ∆[parameter]500, respectively. Patients with an increase in CO after a 500-ml infusion over 15 minutes (∆CO500) of 15% or more were classified as responders (Rs), and those without an increase were classified as non-responders (NRs) . An expeU test. The Friedman test and Wilcoxon rank-sum test were used to detect changes over time  within the same group. Linear correlations were tested by using the Spearman rank correlation and Pearson tests for non-normal and normal distributions, respectively. Receiver operating characteristic (ROC) curves were constructed to evaluate the ability of each parameter to predict fluid responsiveness after VE by varying the discriminating threshold of each measurement. The area under each ROC curve (AUC) was calculated and expressed as AUC ± standard deviation (SD). Cutoff values were chosen with the highest Youden index, calculated as [1 - (1 - sensitivity) - (1 - specificity)]. Probability (P) values of less than 0.05 were considered statistically significant. Data are presented as mean ± SD if normally distributed or as median (interquartile range) if not normally distributed.The SPSS 16.0 software  was used for statistical analyses. Normality was tested by using the Shapiro-Wilk test, and comparisons of hemodynamic parameters between Rs and NRs were assessed by using the Mann–Whitney P = 0.32), LVEF (P = 0.33), and CVP (P = 0.40) were similar between the two groups. Significant differences in the following were predictive of fluid responsiveness: systolic blood pressure (SBP), VTI, SV, and CO. The last three parameters were clearly lower among Rs than NRs at baseline.In total, 55 patients were included; 5 patients were excluded because adequate images could not be obtained. Table P = 0.41) and NRs (P = 0.18).Table r = 0.83; P <0.01)  . The Bland-Altman plot presenting differences between ∆CO50 and ∆CO500 is given in Figure Figure P <0.01), 0.96 ± 0.03 (P <0.01), and 0.95 ± 0.03 (P <0.01), respectively. The AUCs for VTI, SV, and CO at baseline had lower accuracy for predicting fluid responsiveness compared with ∆VTI50 and ∆CO50.As shown in Figure Table Fluid resuscitation is the cornerstone of hypovolemic shock, severe sepsis, or septic shock patient management. However, when patients are on the relatively flat part of the Frank-Starling curve, preload VE can be deleterious. Consequently, monitoring volume responsiveness has become increasingly important in the ICU.P = 0.41) and NRs (P = 0.18). This means that crystalloid distributed rapidly through the extracellular compartment, which did not affect sympathetic stimulation or cardiac reflexes. Our study suggested that there was a significant difference in SBP between Rs and NRs. However, the literature suggests that SBP does not reliably predict a patient’s response to fluid administration, which was affected by drugs, stress, pain, fever, anemia, intrinsic heart disease, and many other factors . With D being constant in a given patient, VTI and SV changes are expected to be equal. Over the past decade, a number of reports have correlated ∆VTI with fluid responsiveness [Recently, dynamic echocardiographic parameters ∆SV, ∆CO, and ∆VTI induced by VE or PLR have gained increased attention; moreover, VE and PLR can reliably predict fluid responsiveness regardless of the ventilation mode or underlying cardiac rhythm in the majority of ICU patients ,27-30. Hsiveness ,29,33-35et al.[As demonstrated by the Frank-Starling curve, the beginning of the fluid challenge induces a more significant increase in stroke volume on the steep portion of the curve because of a preload reserve in the left ventricular function. Muller et al. proposedet al.,37 and tet al.,38.r = 0.83 (P <0.01) and the correlation between ∆CO50 and ∆CO500 was r = 0.90 (P <0.01), respectively. Moreover, Bland-Altman-like analysis of the same data indicated that there was a good concordance for two measurements. This was associated with a trend that a higher VE can lead to a higher change of CO.In the present study, the relatively high correlation coefficient between ∆VTI50 and ∆VTI500 was The ROC analysis of echocardiographic parameters revealed that a response to 50-ml crystalloid bolus over 10 seconds enables accurate prediction of fluid responsiveness, particularly ∆VTI50 and ∆CO50. The AUCs for these echocardiographic parameters were above 0.90. The best AUC cutoff for ∆CO50 when predicting fluid responsiveness was 6% , suggesting that a mini-fluid challenge of 50-ml crystalloid bolus over 10 seconds enables accurate bedside prediction of a 15% increase in ∆CO500 in 27 adults. In Figure This present study has several limitations. First, the patient sample was small and may limit the external validity. Second, because our study excluded patients with cardiac arrhythmias, we do not know its applicability in this setting. Third, transthoracic echocardiography has many specific limitations: It is technically demanding, and many ICUs may lack suitably skilled operators. The sharpness of ultrasound images will be affected by obesity, chest deformity, high abdominal pressure, patient positioning, and other factors. Fourth, we did not assess the impact of measuring time. This assessment deserves further studies.et al. [In contrast to the study by Muller et al. , the pre• Echocardiography offers a convenient, non-invasive, dynamic, and qualitative assessment of volume responsiveness in the ICU.• By using a smaller fluid volume and faster infusion rate than ever before, the study wanted to find a relatively accurate and sensitive fluid challenge technique with more decrease morbidity and mortality.• The variation of CO and VTI measured by transthoracic echocardiography after the administration of 50-ml crystalloid solution over 10 seconds is able to reliably predict fluid responsiveness among critically ill patients.• Despite a lower sensitivity and several limitations, the mini-fluid challenge technique may provide a new research perspective for effective and rapid prediction of fluid responsiveness in the future.∆: Delta; ∆CO50: variation of cardiac output after infusion of 50 ml of fluid over 10 seconds; ∆CO500: variation of cardiac output after infusion of 500 ml of fluid over 15 minutes; ∆SV50: variation of stroke volume after infusion of 50 ml of fluid over 10 seconds; ∆SV500: variation of stroke volume after infusion of 500 ml of fluid over 15 minutes; ∆VTI50: variation of velocity time index after infusion of 50 ml of fluid over 10 seconds; ∆VTI500: variation of velocity time index after infusion of 500 ml of fluid over 15 minutes; AUC: area under the receiver operating characteristic curves; CO: cardiac output; CVP: central venous pressure; D: aortic diameter; HR: heart rate; ICU: intensive care unit; IVC: inferior vena cava; LEDV: left ventricular end-diastolic volume; LESV: left ventricular end-systolic volume; LOA: limit of agreement; LVEF: left ventricular ejection fraction; NR: non-responder; PLR: passive leg raising; R: responder; ROC: receiver operating characteristic; SBP: systolic blood pressure; SD: standard deviation; SV: stroke volume; VE: volume expansion; VTI: aortic velocity time index.The authors declare that they have no competing interests.YW performed the data collection and drafted the manuscript. ZZ performed the statistical analysis and assisted in the drafting of the manuscript. SZ and BL conceived of the study, participated in its design, and coordinated the study. All authors read, revised, and approved the final manuscript.YW, SZ and BL work at the Department of Critical Care Medicine, Affiliated Provincial Hospital of Anhui Medical University. ZZ works at the Department of Critical Care Medicine, Affiliated Hospital of Neurology Institute, Anhui University of Traditional Chinese Medicine.


The optimal management of rectal cancer is achieved through a shared multidisciplinary decision making process with accurate staging by imaging being critical for treatment planning. Good quality, high-resolution MRI has become the imaging gold standard as it allows consistent staging and stratification of patients into distinct prognostic groups according to MR-findings. Imaging features other than T and N have been proven to influence patient outcomes, and increasingly these features are taken into consideration when determining treatment options: distance of tumour to the potential circumferential margin (CRM), presence of tumour within the extramural rectal vessels (EMVI), discontinuous tumour deposits (N1c), relationship to the intersphincteric plane in low rectal tumours and to pelvic compartments in advanced disease. The presence or absence of proven adverse MR features should be included in the MRI report and shared with the patient when treatment choices are offered. MRI enables the identification of high risk tumours where the use of neoadjuvant therapy is justified and is a robust method of identifying patients with a strong likelihood of complete response after preoperative treatment. Imaging plays a major role in treatment decision making of rectal cancer patients. The concept of tumour-spread assessment has markedly changed during the last decade. Rather than just T and N stage, other imaging prognostic factors and imaging-based assessment of the surgical planes can help the colorectal teams make decisions to improve outcomes  is a standard of treatment. The term ‘locally advanced’ and its definition varies between centres and countries. The avoidance of preoperative therapy in patients with a low risk of local recurrence or distant failure is gaining widespread acceptance in Europe but is based on outcome data that relates to audited surgical total mesorectal excision (TME) with removal of tumour and draining nodes in a single ‘package’. There is also widespread agreement that regardless of T and N stage, tumour extending to within 1 mm of the surgical TME plane  is associated with a high risk of local recurrence. Such patients are accurately identified by MRI and offered preoperative chemoradiotherapy –4. MRI fStage-directed treatment plan based on pretreatment staging of rectal cancer is recommended by the National Comprehensive Cancer Network (NCCN) guidelines . ClinicaThe depth of spread beyond the muscularis in millimetres has been proven to influence survival rates and is considered as an important prognostic factor that enables precise prognostic stratification within the T3 subcategory , 14. Basp < 0.0001) and a significantly lower rate of CRM involvement 4 % versus 20 % in favour of preoperative chemotherapy [Stratification of colon cancer according to the extramural spread beyond the muscularis propria has been proven to be useful method for identifying high risk patients who could benefit from neoadjuvant chemotherapy . In a fuotherapy .Whatever the depth of tumour infiltration at the level of infiltrating border assessment of the whole mesorectum is crucially important. Relationship of the tumour to the mesorectal fascia should always be reported. A distance on MRI of less than 1 mm to the mesorectal fascia was proven to predict pCRM status Fig.  25]; mo; mo25]; The mesorectum should also be assessed for presence of suspicious tumour deposits and extravascular tumour invasion; for tumours arising at a height of <6 cm from the anal verge, MRI detection of EMVI is independently associated with a risk of pathologic CRM involvement ••.Published data suggest that morphological features such as heterogeneous signal intensity and lymph node irregular capsule border are accurate predictors of metastatic spread within the nodes , 28. HowSome nodular structures could represent vascular deposits, a phenomenon that is difficult to assess on final histology as absence of preserved nodular capsule preclude differentiating malignant lymph node with affected capsule from tumour deposit. However, the latter predicts poorer prognosis , 33. Higp < 0.0001) in those with no changes of EMVI status [Both MRI and histopathologic EMVI are strong predictors of poor prognosis, particularly as a predictor of tumour metastasising to the liver . In multI status . SurvivaI status •• and itI status .Mucinous adenocarcinomas are associated with worse disease-free and overall survival outcomes compared to non-mucinous tumours and appear to be less sensitive to preoperative CRT when compared with non-mucinous tumours , 38. HisDifferent MRI techniques have been proposed as a tool for tumour response assessment; one of these is modified mr RECIST which is based on objective measurements of the change in craniocaudal length of measurable disease. According to new guidelines to evaluate the response to treatment in solid tumours, at least 30 % reduction rate should be considered as good or favourable response . HoweverAn MR regression-grading system based on the principles of the modified Mandard TRG system has been proposed for assessment of tumour response in rectal cancer classifying post-treatment changes. This divides response into five categories based on the proportion of intermediate tumour signal intensity versus low fibrosis signal intensity within the treated tumour  local excision could be considered as the main treatment option.Since the 1990s, endorectal ultrasound (ERUS) has been considered as the best diagnostic modality for early rectal cancer, and accuracy rates were reported to be as high as 90 % –50. Howep < 0.01). The presence of at least 1 mm of submucosal layer identified as high signal intensity line between the tumour-advancing edge and muscularis suggest that LE is safe and feasible to perform  [In patients with CRM involvement, R0 resection can only be achieved if surgical planes are extended beyond the TME plane, and the radiologist plays a fundamental role in both identifying these cases and correctly assessing the surgical planes required for histological clearance. High-resolution T2-WI MR allows identifying surgical planes and indicates when extralevator abdominoperineal resections (ELAPE) and exenterative surgery could potentially be performed . On mult= 0.002) .A useful system is to divide the pelvis into six surgical compartments /sciatic nerve/sacral nerve branches (S1/2); laterally: pelvic fascia/pelvic sidewall/internal/external iliac vessels/sacrotuberous/sacrospinous ligaments/piriformis/obturator internus muscles; infralevator: Levator muscles/sphincter complex; anterior urogenital triangle/perineum: vaginal introitus/urethra/retropubic space)  Fig. 1i1i.Treatment decisions are no longer made by a single specialist but a colorectal unit team, when pre-treatment tumour spread assessment drives the type of treatment to be chosen. Imaging plays a pivotal role in guiding the treatment management. Surgeons push forward radiologists to improve staging accuracy of most accepted tumour spread criteria such as T and N; however, the paradigm of tumour spread prognostic factors has shifted and new imaging biomarkers are emerging into the routine radiological practise.Such tumour spread markers as EMVI, relationship to the mesorectal fascia and intersphincteric plane (CRM status), presence of mucin component within the tumour have a greater influence on patient outcomes than T and stage and therefore should now be factored into treatment decisions.Imaging enables better stratification of patients in risk groups for local and distal failure, helps to clarify safe surgical planes, identifies those patients who could potentially benefit from preoperative treatment and ultimately could identify those patients for whom surgery may be safely omitted of deferred.


The thentest design aims to detect and control for recalibration response shift. This design assumes (1) more consistency in the content of the cognitive processes underlying patients’ quality of life (QoL) between posttest and thentest assessments than between posttest and pretest assessments; and (2) consistency in the time frame and description of functioning referenced at pretest and thentest. Our objective is to utilize cognitive interviewing to qualitatively examine both assumptions.We conducted think-aloud interviews with 24 patients with cancer prior to and after radiotherapy to elicit cognitive processes underlying their assessment of seven EORTC QLQ-C30 items at pretest, posttest and thentest. We used an analytic scheme based on the cognitive process models of Tourangeau et al. and Rapkin and Schwartz that yielded five cognitive processes. We subsequently used this input for quantitative analysis of count data.Contrary to expectation, the number of dissimilar cognitive processes between posttest and thentest was generally larger than between pretest and posttest across patients. Further, patients considered a range of time frames when answering the thentest questions. Moreover, patients’ description at the thentest of their pretest functioning was often not similar to that which was noted at pretest. Items referring to trouble taking a short walk, overall health and QoL were most often violating the assumptions.Both assumptions underlying the thentest design appear not to be supported by the patients’ cognitive processes. Replacing the conventional pretest–posttest design with the thentest design may simply be replacing one set of biases with another.The online version of this article (doi:10.1007/s11136-015-1175-4) contains supplementary material, which is available to authorized users. Change in patients’ quality of life (QoL) is most commonly assessed by means of the prospective baseline and follow-up design, also known as the pretest–posttest design –5. HowevThe retrospective pretest–posttest design is a commonly used approach to detect and control for recalibration response shift when measuring change in QoL –14. The To the best of our knowledge, only Westerman et al. , 19 haveTo examine the cognitive processes underlying the responses of patients with cancer to QoL (thentest) items, we have developed an analytic scheme  based onIn a prior study, we used this analytic scheme to examine the content of the cognitive processes underlying patients’ responses to QoL items in a pretest–posttest design to study the assumption of consistency in the content of respondents’ cognitive processes over time. Our results showed that the content of all cognitive processes changed over time .The present study builds on these prior results by examining whether the content of each distinct cognitive process underlying patients’ responses remains similar or changes between posttest and thentest. Since the thentest design is assumed to control for inconsistencies in respondents’ QoL assessment between pretest and posttest, we expect more congruence in the content of the cognitive processes underlying responses to the posttest and thentest than to the pretest and posttest. To comprehensively address both assumptions underlying the thentest design, we will also examine whether patients accurately recall their pretest functioning when completing thentest items. Finally, we will examine these assumptions at the individual patient and item level.n = 50) [This study continues work from our prior research project where pretest and posttest assessments (n = 50)  were eitn = 50) . In the n = 50)  and hereTo include a heterogeneous sample with variation in cognitive processes used in arriving at responses to QoL (thentest) items, we purposefully selected patients with cancer with respect to gender, age and tumor site. All patients were undergoing radiotherapy at the Department of Radiation Oncology at the Academic Medical Center (AMC) in Amsterdam.In accordance with designs commonly used in treatment evaluation, pretest assessments were administered prior to, and posttest and thentest assessments at the end of a salient health-related intervention. Pretest interviews took place on the same day the patient had an appointment for the CT-simulator to prepare the treatment or received the first radiation treatment. The posttest and thentest interviews were conducted on patients’ last day of radiotherapy. The interviews were conducted at the Department of Radiation Oncology of the AMC in 2008 by two researchers  not involved in the patient’s clinical care. All interviews were audio-recorded and transcribed verbatim.To limit patient burden, we used seven items from the 30-item EORTC QLQ-C30 , a HRQoLWe used Hak et al’s  Three-Stresponse, but rather to provide a renewed judgment. We used non-leading probes for the thentest items, such as “You just referred to your functioning prior to the start of radiotherapy, could you explain to me how you tried to recall this period?”After completion of the posttest think-aloud interview, we introduced the thentest by jogging the patient’s memory about the time point at which s/he conducted the pretest think-aloud interview. As recommended, we tried to prompt recollection of the day and time of the pretest interview and provided cues to elicit patient’s memory , 28. We Qualitative content analysis of all interviews was independently carried out by two researchers  using MAXqda software . The proThe second assumption of accurate recall underlying the thentest was operationalized by examining whether the time frame employed and the description of pretest functioning provided in answering each thentest item were similar to those of the corresponding pretest item. To that end, we compared the content of the cognitive processes comprehension/frame of reference and retrieval/sampling strategy at pretest and thentest. The codes were used as input for quantitative analyses. Again, all findings were discussed and consensus was negotiated in case of differences. Throughout the period of data collection and analysis, all codes and subsequent analyses were also discussed with co-authors FvZ and MS. Field notes were taken of decisions made to ensure consistency in the analysis.Of the 38 eligible patients approached, 12 choose not to participate explaining they considered it too burdensome to be cognitively interviewed prior to and at the end of radiotherapy. Twenty-six patients gave written informed consent. One patient was unable to participate in the posttest and thentest interviews due to severe health deterioration, and one patient could not be interviewed at the end of radiotherapy due to logistical problems. The median number of days between the pretest interview, and the posttest and thentest interviews was 46 days . Table Table At the patient level, there appears to be a linear trend such that patients with more discrepancies in cognitive processes between posttest and thentest also have more discrepancies in cognitive processes between pretest and posttest  and 7  induced the largest discrepancy in cognitive processes between the posttest–thentest versus pretest–posttest comparison  similarity in time frame and description of pretest functioning; (2) similarity in time frame and dissimilarity in description of pretest functioning; 3) dissimilarity in time frame and similarity in description of pretest functioning; and (4) dissimilarity in time frame and dissimilarity in description of pretest functioning. Whereas category 1 supports the assumption, the remaining three do not. The number of items across patients that was found to support the assumption was 60, whereas those in the three deviant categories totaled 102. The patients in this study thus more often used other time frames than those intended by the researchers, and provided a different description at the thentest than at the pretest. No clear pattern could be discerned regarding the four individual and two aggregated categories at the patient level. Table  dissimilIn this study, we examined the two assumptions inherent to the thentest design. Contrary to our expectations, the number of dissimilar cognitive processes between posttest and thentest was generally larger than between pretest and posttest across patients. Thus, these findings do not support the thentest design’s first assumption that the administration of the posttest and thentest in temporal proximity would induce a comparable content in cognitive processes. We also examined the thentest’s second assumption of accurate recall of pretest functioning. In comparison with their responses at pretest, patients more often provided a different description of their initial functioning and referred to a different time frame when responding to the thentest. In conclusion, both assumptions underlying the thentest design appear not to be supported by the cognitive processes described by the patients in this sample.At the item level, items 1 , 6  and 7  were found to most notably violate the assumptions. Perhaps these items leave most room for personal interpretation . However, as we can imagine that other items  may also invite different views over time, it remains uncertain as to why some items seem to violate the assumptions more than others in this study. No discernable patterns were found at the patient level other than that patients with more discrepancies in the content of the cognitive processes between posttest and thentest also displayed more discrepancies between pretest and posttest.According to Rapkin and Schwartz , change Several studies have examined the accuracy of retrospective assessment of QoL as opposed to its prospective assessment. Some of these studies advocate the use of the thentest for treatment evaluation and consider it a more valid approach to measure change than standard, prospective measurement , 31–33. To increase our insight into patients’ response strategies, future studies are needed that examine how and why patients arrive at their answers. For example, studies could be designed in which patients are confronted with their pretest scores, after they have completed the thentest. Patients could subsequently be invited to comment their scores. Such interviews might provide valuable insight into patients’ own explanations of inconsistencies in the time frames employed and the descriptions of pretest functioning provided. In addition, to further elucidate actual response shift processes, it would be interesting to compare the content of all five cognitive processes between pretest and thentest in a more open and explanatory qualitative approach.A number of limitations of this study warrant attention. First, given the small number of patients and the purposeful sampling strategy, we do not know how representative this patient group is and to what extent the characteristics of this particular sample may have affected the results. We do know, however, that patients who refused participation found the study too burdensome, which might indicate that the most severely ill patients were not included in this study’s sample. The interpretation of our findings needs thus be limited to the less severely ill . Second,We would also like to put forward a number of considerations when interpreting the results of this study. First, as is common in employing a thentest, we first have tried to revive patients’ memory about their pretest functioning. The thentest is most commonly administered in a written format. However, in the present study, we have orally instructed patients to think back to their pretest functioning. It is plausible that an oral instruction and subsequent think-aloud interview might have increased patients’ efforts to recall their pretest functioning. Consequently, in the context of conventional treatment evaluation, the results may show more deviation between pretest and thentest than was the case in the current study.Second, our focus on the number of cognitive processes that has changed does not imply that a quantitative change is more important than a change in the content of cognitive processes. Dependent on the context, a change in one cognitive process may render responses over time more incompatible than changes in two or more cognitive processes. However, since the thentest design assumes that the content of each distinct cognitive process does not change between posttest and thentest, using the number of changed cognitive processes in this sample is informative. However, the absolute numbers should be interpreted with caution.prior to her cancer diagnosis and treatment, and her thentest QoL—referred to the period after diagnosis but prior to treatment—in reference to the QoL of life at the posttest. We concluded that she adopted a different standard of comparison at posttest than at thentest. However, one may argue that she is doing exactly what we want her to do at the thentest: judging her former QoL as compared to her current QoL. It should be noted that we adopted the same strict criteria for testing the first assumption for the conventional pretest–posttest design [Third, the strict operationalizations of the two assumptions adopted here naturally affect the results. Both the pretest and thentest instruct patients to rate their QoL in the week prior to the pretest interview . For example, for the 59-year-old female with gynecological cancer Table  the stant design . TherefoFinally, our interpretation of recall bias merits attention as inconsistent time frames between pretest and thentest are not always indicative of recall bias per se. In the instances where patients did not adopt the requested time frame at the pretest, reference to the week prior to the pretest at the thentest (indication of accurate time frame) would still be counted as a discrepancy. Whereas the interpretation of recall bias in those instances is disputable , they stIn conclusion, the cognitive processes underlying patients’ responses to thentest items in this study appear not to be in line with the assumptions of (1) consistency in the content of the cognitive processes underlying responses to the posttest and thentest, and (2) accurate recall of pretest functioning. Rather, our data suggest that these patients select personally meaningful time frames and content when (retrospectively) assessing their QoL, which might deviate from the time frames considered relevant by researchers. The question arises whether the thentest is a suitable alternative in controlling for inconsistencies in respondents’ cognitive processes over time. However, as argued previously , 39, retSupplementary material 1 (DOCX 30 kb)Below is the link to the electronic supplementary material.


KIT or PDGFRA mutations. In this study we characterize a newly established GIST xenograft model, UZLX-GIST9, and evaluate the in vivo response of the model to standard TKIs .Acquired resistance to tyrosine kinase inhibitors (TKIs) in gastrointestinal stromal tumours (GISTs) is most commonly caused by secondary Tumour fragments from a metastatic lesion of a GIST patient clinically progressing after treatment with imatinib, sunitinib and regorafenib were engrafted in a nude, immunodeficient mouse. Upon sequential passaging from mouse to mouse, tumour fragments were collected for histopathological and molecular characterization. The sensitivity of the model to treatment with TKIs was evaluated in 28 mice . Mice were grouped as follows: control (untreated), imatinib (50 mg/kg/BID), imatinib (100 mg/kg/BID), sunitinib (40 mg/kg/QD), and regorafenib (30 mg/kg/QD). After three weeks of oral treatment, tumours were collected for subsequent analysis. The efficacy of treatment was assessed by tumour volume, histopathology and Western immunoblotting.KIT mutational profile (p.P577del + W557LfsX5+ D820G) remains the same as the original tissue sample originating from an intraspinal metastatic site. Three week treatment with different TKIs showed that the model is resistant to imatinib. Sunitinib induces tumour growth delay and regorafenib reduces the tumour burden by 30% as compared to control animals. While none of the TKIs had a significant effect on cell proliferation or cell survival, a remarkable increase of necrosis and significant reduction of microvessel density was observed under sunitinib and regorafenib. Western immunoblotting showed a mild reduction in KIT and AKT activation only in regorafenib treated tumours.UZLX-GIST9 maintains the same typical morphological features and immunohistochemical characteristics as the original patient biopsy and expresses CD117 and DOG1. The KIT exon 11 and 17 mutations and maintaining the pheno-and genotype of the original tumour. UZLX-GIST9 shows different levels of response to standard TKIs. This model will help to study TKI resistance and to explore novel treatment approaches for patients with TKI-resistant GIST.We established a novel human GIST xenograft, UZLX-GIST9, harbouring  KIT or the PDGFRA  gene are the main oncogenic drivers as the reference sequence were subcutaneously engrafted in both flanks with UZLX-GIST9 and were used in two subsequent experiments. A detailed overview of treatment groups and schedules can be found in Table For the ge 4 n = ] were suFixed tumour specimens were embedded in paraffin blocks and cut into 4 μm sections on the Microm HM 340 E microtome for haematoxylin and eosin staining (H & E) or IHC. Histological response (HR) was graded by assessing the magnitude of necrosis, fibrosis and myxoid degeneration: grade 1 (0%–10%), grade 2 (>10% and ≤50%), grade 3 (>50% and ≤90%), and grade 4 (>90%) as described before,25. AsseFor Western immunoblotting, tumour lysates were prepared from snap frozen tumour specimen as described previously. Chemiluversus the last day of the experiment. The comparison between different treatment groups was performed using the Mann–Whitney U (MWU) test. The statistical significance level was defined as p < 0.05 and the STATISTICA software  was used for all calculations.Wilcoxon’s matched paired (WMP) test was used to compare tumour volumes on day 0 (baseline) Since April 2012, the UZLX-GIST9 xenograft model has been maintained by sequential transplantations in NMRI nude mice and currently reached the 7th passage. Histopathological evaluation showed a similar morphological appearance of the ex-mouse UZLX-GIST9 tumours as compared to the original intraspinal GIST metastatic sample obtained from the patient. Microscopic analysis revealed a predominantly spindle cell morphology and diffuse CD117 (KIT) immunoreactivity in the original GIST metastasis and in ex-mouse tumours  and exon 17 (p.D820G) mutations as found in the original GIST intraspinal metastasis which was used for engraftment. The mutational profile remained stable throughout the different passages in the mice.The mutational analysis of UZLX-GIST9 xenografts revealed the same in vivo. In general, the tumour burden increased steadily in untreated and imatinib treated cohorts, showing a relative increase to 282, 278, and 238% for untreated, imatinib 50 mg/kg/BID and imatinib 100 mg/kg/BID treated animals as compared to baseline   was assessed by evaluating the magnitude of necrosis, fibrosis and myxoid degeneration on H & E stainings of tumours collected at the end of the Untreated UZLX-GIST9 tumours are characterized by brisk mitotic activity (39 mitotic figures/10 HPF on average) on H & E staining. Neither TKI treatment induced a reduction in mitotic activity; rather a slight increase of mitotic activity was observed under TKI treatment, which was even statistically significant in the regorafenib treated cohort Table . No signMVD analysis showed that none of the imatinib regiments significantly lowered vessel density as compared to untreated control tumours. In contrast, both sunitinib and regorafenib yielded a 1.4 reduction of microvessel density, which was significant when compared to control  Table .By Western immunoblotting we observed expression of KIT and its main signalling mediators in all untreated UZLX-GIST9 tumours Figure . In genein vitro and in vivo models are required to test new agents and assess the molecular mechanism of action of innovative drugs.Imatinib treatment in GIST is a paradigm for the use of targeted treatments in solid tumours. Nevertheless, despite the remarkable effects of this and other TKIs in GIST, with time the vast majority of patients will inevitably develop resistance against these agents. Importain vivo experiments[KIT exon 17 mutation[To date, the number of GIST xenograft models available for research is rather limited. A few xenografts have been established upon subcutaneous injection of GIST-T1, GIST48, GIST430, GIST882 or GIST882Ly cell lines,22,27-29eriments,27. Onlyeriments-23,30-32mutation.Here we are presenting data from a novel GIST xenograft model (UZLX-GIST9) based on a tumour biopsy derived from a patient who had been treated for almost 10 years with a variety of TKIs, including imatinib, sunitinib, regorafenib and nilotinib. To date, the new model is maintained for over two years and 7 passages and continues to grow.KIT exon 11 and exon 17 mutations , suggesting resistance to commonly used TKIs.UZLX-GIST9 mimics the morphological, immunohistochemical and molecular features of the original intraspinal GIST metastasis with spindle cell morphology, diffuse CD117 and DOG1 immunoreactivity. The mutational analysis of the xenograft shows Indeed, we could show imatinib resistance in the dose range tested. Conversely, the model still showed some level of response to other TKIs (sunitinib and regorafenib), even though we did not observe any anti-mitotic or pro-apoptotic effects of the tested agents. Sunitinib yielded tumour growth delay and regorafenib induced some regression in tumour volume. Most likely, these effects on tumour burden are related to the prominent induction of necrosis observed under regorafenib and sunitinib, which was not seen under imatinib treatment. In addition, only in regorafenib treated tumours a slight inhibition of KIT and AKT activation was observed. The mild increase in mitotic activity in the viable tumour areas under regorafenib treatment is most likely the result of selective pressure by the high degree of necrosis observed in these tumours and the multi-targeted nature of the TKI, resulting in the selection of the most aggressive and mitotically active tumour cells.These findings provide evidence for the characterization of UZLX-GIST9 as a truly imatinib-resistant model. Intriguingly, despite the lack of a seemingly direct inhibitory effect on cell proliferation and survival we observed a decrease in tumour growth rate or even tumour volume reduction for sunitinib and regorafenib, respectively. Both drugs are known to have anti-angiogenic effects, and we have shown that both drugs significantly reduced MVD in the newly established GIST xenograft model when compared to untreated control tumours,15. We tKIT exon 11 and exon 17 mutations has been established. This novel xenograft model resembles the morphological and molecular features of the original tumour that had failed treatment with four different potent TKIs, including the three approved agents for the therapy of advanced, metastatic GIST. The model exhibits strong imatinib resistance; tumour growth and high tumour cell proliferation was observed under sunitinib. Regorafenib shows significant anti-tumour effects in this model, which seem independent of KIT inhibition. Furthermore, both sunitinib and regorafenib significantly lower MVD and induce necrosis in the UZLX-GIST9 model. Overall, we present a novel GIST research tool which will be very valuable for the development of novel experimental treatment approaches and hopefully translate into direct patient benefit.In summary, a novel human GIST xenograft with The patient agreed with using a tissue sample collected during routine surgery for research purposes. She could not give written informed consent for the current publication as she died in August 2012.The authors declare that they have no competing interests.Histomorphology , molecular analysis , data collection , analysis of data , project coordination , and manuscript preparation . All authors read and approved the final manuscript.


In this paper, the performance of an individual aiming at guiding a self-organized group is numerically investigated. A collective behavioural model is adopted, accounting for the mutual repulsion, attraction and orientation experienced by the individuals. Moreover, these represent a set of solid particles which are supposed to be immersed in a fictitious viscous fluid. In particular, the lattice Boltzmann and Immersed boundary methods are used to predict the fluid dynamics, whereas the effect of the hydrodynamic forces on particles is accounted for by solving the equation of the solid motion through the time discontinuous Galerkin scheme. Numerical simulations are carried out by involving the individuals in a dichotomous process. On the one hand, an aspirant leader (AL) additional individual is added to the system. AL is forced to move along a prescribed direction which intersects the group. On the other hand, these tend to depart from an obstacle represented by a rotating lamina which is placed in the fluid domain. A numerical campaign is carried out by varying the fluid viscosity and, as a consequence, the hydrodynamic field. Moreover, scenarios characterized by different values of the size of the group are investigated. In order to estimate the AL's performance, a proper parameter is introduced, depending on the number of individuals following AL. Present findings show that the sole collective behavioural equations are insufficient to predict the AL's performance, since the motion is drastically affected by the presence of the surrounding fluid. With respect to the existing literature, the proposed numerical model is enriched by accounting for the presence of the encompassing fluid, thus computing the hydrodynamic forces arising when the individuals move. Social interactions involve the decision processes emerging in many animal groups, such as flocks of birds, schools of fishes or swarms of insects social approaches, the prediction of the leader identification and decision making processes emerging in a set of self-organized individuals is enriched by accounting for the presence of the encompassing fluid. Specifically, the lattice Boltzmann (LB) method is adopted to predict the fluid dynamics In this paper, beside the uniquely The rest of the paper is organized as follows. In the Here, the adopted numerical methods are briefly recalled and the proposed algorithm of computation is presented.According to Making reference to Concerning the presence of an obstacle (e.g. a rotating lamina), it is identified as an additional set of individuals with respect to the group is repulsed, whereas the attraction-orientation rule is neglected. Such tendency is enforced even for AL.The two-dimensional Bhatnagar-Gross-Krook In order to account for the presence of an immersed body (orange circles) in the fluid lattice background at the position In order to compute the time- and force-dependent displacements The above mentioned numerical methods are combined in a proper strategy, consisting of an algorithmical sequence of actions which are sketched in The problem set-up is sketched in In A deeper investigation on the space-time evolution of the group is carried out by sketching the spatial configuration of the individuals at ered see , resultshttps://www.youtube.com/playlist?list=PL-z4SoMtQxJSHsrIKoZDimokoKf6klFFJ).Finally, notice that the author creates a website where the animations showing the velocity field and spatial configuration of the individuals are present (Previous simulations have been carried out from a deterministic point of view, by neglecting the presence of disorder and disturbance. As known, the dynamics of a group of self-organized individuals can be affected by several random conditions, such as the initial position or the orientation of the individuals. Here, the scenario characterized by In this paper, the collective dynamics of a group of self-organized individuals immersed in a fictitious viscous fluid has been investigated. The group interacts with an additional individual, AL, which aims at becoming a leader. The effect of the fluid forces has been elucidated by combining the LB-IB-TDG strategy for fluid and solid dynamics to the collective behavioural equations. It has been found that the fluid tends to create a more compact group, thus enforcing a deeper exchange of information among the individuals. As a consequence, AL can successfully guide the individuals. In addition, groups characterized by progressively larger size have been involved in the computations, showing a splitting effect which increases with


To determine the diagnostic performance of DWI volumetry for the assessment of complete response after CRT in patients with locally advanced rectal cancer and to compare with volumetry on standard T2- weight MRI, by means of volumetric signal intensity measurements.We retrospectively analyzed 25 patients with locally advanced rectal cancer. Patients underwent pre and post-CRT standard T2 weight MRI and DWI MRI. We placed free-hand regions of interest and each tumour-containing section to determine pre and post-CRT tumour volumes.Histologic findings were the standard of reference.Volumetry of the viable tumour remmants based on signal intensity characteristic on DW images after CRT was shown to be more valuable than volumetry of the tumour based on morphologic T2w images for the differentiation between a pCR and residual tumour in locally advanced rectal cancer.


Patient stratification for therapeutic response mainly focus on biomarker expression on cancer cells. Contrary, the tumour micromileu is less considered, although stromal cells and leukocytes play a crucial role in tumour progression and drug resistance. The present study investigates the impact of tumour infiltrating lymphocytes on catumaxomab efficacy in CRC.Fresh tumour samples from 27 CRC patients were used to prepare tumour spheroids. Autologous PBMCs were isolated by ficoll density gradient. Tumour spheroids co-cultured with or without PBMCs were treated with catumaxomab for 96h. The cellular composition of the spheroids and the therapeutic impact on epithelial cells and leukocytes were measured by FACS analysis. The metabolic activity of the co-cultures was determined by the ATP assay.Similar to their original cancers all spheroid models consisted of a sufficient high fraction of EpCAM positive tumour cells indicating CRC an appropriate cancer type for catumaxomab therapy. In contrast, the fraction of CD45 positive leukocytes in spheroids was low mimicking the parental tumours. The effector cell to target cell  ratio ranged from 0,05:1 to 1,92:1. Treatment of these spheroids with catumaxomab revealed no significant therapeutic effect. Addition of patient specific PBMCs changed the E:T ratio up to 7:1 and had a significant impact on antibody efficacy. Catumaxomab induced cell death was found up to 52%  depending on the individual E:T ratio. After catumaxomab treatment co-cultures demonstrated metabolic stimulation underlining the functionality of the leukocytes.A dual biomarker system is required to select appropriate CRC patients for catumaxomab treatment. EpCAM expression on cancer cells is required, but the leukocyte infiltrate is the key predictor for the therapeutic response of catumaxomab. Both extent and functionality of the tumour infiltrating leukocytes have to be determined in the individual tumour sample for targeted therapy using the trifunctional monoclonal antibody.