Development and validation of an ELISA to measure regenerating island-derived protein 3E in canine blood.

BACKGROUND: Regenerating island-derived proteins (REG) are upregulated in people with sepsis, pancreatitis, and gastrointestinal diseases. One member of the REG family, namely REG3E, was recently identified in pancreatic tissue and plasma of dogs, with high expression in pancreatitis and sepsis. OBJECTIVES: We aimed to develop and validate an ELISA to measure REG3E concentrations in canine blood. METHODS: An indirect sandwich ELISA was developed using recombinant canine REG3E protein and polyclonal anti-canine REG3E antibodies raised in guinea pigs and rabbits. Antibody specificity was assessed using western blot and mass spectrometric analysis of protein purified from canine plasma. Assay validation included evaluation of dilutional linearity, parallelism, spiking recovery, repeatability and reproducibility, stability, interferences, and comparison of serum and heparinized plasma. RESULTS: Antibodies bound specifically to REG3E with no evidence of cross-reactivity with other proteins. The limit of detection of the ELISA was 15 ng/mL, and the lower limit of quantification was 30 ng/mL. The assay demonstrated good to excellent linearity, dilutional and mixing parallelism, and recovery, with mean observed-to-expected ratios of 104%, 107%, 102%, and 92%, respectively, and no evidence of a hook effect. Coefficients of variation were ≤8.5% for repeatability and ≤14.3% for reproducibility at three different levels. Measurements of REG3E in plasma were not significantly influenced by different storage conditions, freeze-thawing cycles, hemolysis, lipemia, or icterus. There was no significant difference between REG3E concentrations in heparinized plasma and serum samples. CONCLUSIONS: The canine REG3E ELISA has acceptable precision, accuracy, linearity, and reproducibility for the measurement of REG3E in canine plasma and serum.

Discriminative performance of pancreatic stone protein in predicting ICU mortality and infection severity in adult patients with infection: a systematic review and individual patient level meta-analysis.

BACKGROUND: Several studies suggested pancreatic stone protein (PSP) as a promising biomarker to predict mortality among patients with severe infection. The objective of the study was to evaluate the performance of PSP in predicting intensive care unit (ICU) mortality and infection severity among critically ill adults admitted to the hospital for infection. METHODS: A systematic search across Cochrane Central Register of Controlled Trials and MEDLINE databases (1966 to February 2022) for studies on PSP published in English using 'pancreatic stone protein', 'PSP', 'regenerative protein', 'lithostatin' combined with 'infection' and 'sepsis' found 46 records. The search was restricted to the five trials that measured PSP using the enzyme-linked immunosorbent assay technique (ELISA). We used Bayesian hierarchical regression models for pooled estimates and to predict mortality or disease severity using PSP, C-Reactive Protein (CRP) and procalcitonin (PCT) as main predictor. We used statistical discriminative measures, such as the area under the receiver operating characteristic curve (AUC) and classification plots. RESULTS: Among the 678 patients included, the pooled ICU mortality was 17.8% (95% prediction interval 4.1% to 54.6%) with a between-study heterogeneity (I-squared 87%). PSP was strongly associated with ICU mortality (OR = 2.7, 95% credible interval (CrI) [1.3-6.0] per one standard deviation increase; age, gender and sepsis severity adjusted OR = 1.5, 95% CrI [0.98-2.8]). The AUC was 0.69 for PSP 95% confidence interval (CI) [0.64-0.74], 0.61 [0.56-0.66] for PCT and 0.52 [0.47-0.57] for CRP. The sensitivity was 0.96, 0.52, 0.30 for risk thresholds 0.1, 0.2 and 0.3; respective false positive rate values were 0.84, 0.25, 0.10. CONCLUSIONS: We found that PSP showed a very good discriminative ability for both investigated study endpoints ICU mortality and infection severity; better in comparison to CRP, similar to PCT. Combinations of biomarkers did not improve their predictive ability.

Early Postoperative Serum Phosphate Drop Predicts Sufficient Hypertrophy After Liver Surgery.

OBJECTIVE: The aim of this study was to assess the impact of postoperative hypophosphatemia on liver regeneration after major liver surgery in the scenario of Associating Liver Partition with Portal vein ligation for Staged hepatectomy (ALPPS) and living liver donation (LLD). BACKGROUND: Hypophosphatemia has been described to reflect the metabolic demands of regenerating hepatocytes. Both ALPPS and LLD are characterized by an exceptionally strong liver regeneration and may be of particular interest in the context of posthepatectomy hypophosphatemia. METHODS: Serum phosphate changes within the first 7 postoperative days after ALPPS (n=61) and LLD (n=54) were prospectively assessed and correlated with standardized volumetry after 1 week. In a translational approach, postoperative phosphate changes were investigated in mice and in vitro . RESULTS: After ALPPS stage 1 and LLD, serum phosphate levels significantly dropped from a preoperative median of 1.08 mmol/L [interquartile range (IQR) 0.92-1.23] and 1.07 mmol/L (IQR 0.91-1.21) to a postoperative median nadir of 0.68 and 0.52 mmol/L, respectively. A pronounced phosphate drop correlated well with increased liver hypertrophy ( P <0.001). Patients with a low drop of phosphate showed a higher incidence of posthepatectomy liver failure after ALPPS (7% vs 31%, P =0.041). Like in humans, phosphate drop correlated significantly with degree of hypertrophy in murine ALPPS and hepatectomy models ( P <0.001). Blocking phosphate transporter (Slc20a1) inhibited cellular phosphate uptake and hepatocyte proliferation in vitro. CONCLUSION: Phosphate drop after hepatectomy is a direct surrogate marker for liver hypertrophy. Perioperative implementation of serum phosphate analysis has the potential to detect patients with insufficient regenerative capacity at an early stage.

The Role of Pancreatic Stone Protein (PSP) as a Biomarker of Pregnancy-Related Diseases.

Background: Pancreatic stone protein (PSP) is a biochemical serum marker that contains levels that are elevated in various inflammatory and infectious diseases. The role of PSP in the diagnosis of these diseases seems to be more important compared to clinically established biochemical serum markers in discriminating the severity of the same diseases. Standard values for PSP in pregnant women in relation to gestational age have been reported recently. Additionally, increased PSP levels have been observed to be associated with renal dysfunction in pregnant women. The aim of this study is to evaluate the diagnostic role of PSP in pregnancy-related diseases, such as pre-eclampsia (PE), hemolysis-elevated liver enzymes, and low platelet (HELLP) syndrome. In addition, the study aims to assess its diagnostic role in inflammation-triggered diseases as preterm premature rupture of membranes (PPROM) or COVID-19-positive pregnant women. Materials and Methods: In this single-centred prospective study performed at a tertiary university hospital between 2013 and 2021, we included 152 pregnant women who were diagnosed with either PE, HELLP syndrome, or PPROM. In December 2020, in the context of the COVID-19 pandemic, the Independent Ethics Committee (IEC) approved an amendment to the study protocol. Depending on the underlying disease, single or serial-serum PSP measurements were assessed. These PSP values were compared to PSP levels of women with normal pregnancies. Results: Pregnant women diagnosed with pre-eclampsia or HELLP syndrome had significantly increased PSP values (mean 9.8 ng/mL, SD 2.6) compared to healthy singleton pregnant women (mean 7.9 ng/mL, SD 2.6, p ≤ 0.001). There was no difference in serum PSP in pregnant women with PPROM compared to women with uncomplicated singleton pregnancies (mean in PPROM: 7.9 ng/mL; SD 2.9 versus mean in healthy pregnancies: 7.9 ng/mL; SD 2.6, p = 0.98). Furthermore, no difference in the PSP values in women with or without intra-amniotic infection was observed (infection: mean 7.9 ng/mL; SD 2.8 versus no infection: mean 7.8 ng/mL; SD 3, p = 0.85). The mean value of PSP in COVID-19-infected women during pregnancy (8.5 ng/mL, SD 2.3) was comparable to healthy singleton pregnancies (mean 7.9 ng/mL, SD 2.6), p = 0.24. Conclusions: The novel serum biomarker PSP is significantly upregulated in pregnant women with pre-eclampsia and HELLP syndrome. Our observations call for the further evaluation of PSP in randomized controlled clinical trials to demonstrate the actual role of PSP in pregnancy-related diseases and whether it may provide new approaches for the management and discrimination of the severity of these gestational conditions.

Serum Pancreatic Stone Protein Reference Values in Healthy Pregnant Women: A Prospective Cohort Study.

BACKGROUND: In non-pregnant populations, pancreatic stone protein (PSP) has been reported to have a higher diagnostic performance for identifying severe inflammatory and infectious disease than other established biomarkers. OBJECTIVE: To generate reference values for serum PSP in pregnancy and compare them to the values of the general healthy population. DESIGN: A prospective cohort study. SETTING: A single center. POPULATION: Healthy women with singleton and multiple pregnancies. METHODS: This is a prospective single-center cohort study. Between 2013 and 2021, samples of 5 mL peripheral blood were drawn from 440 healthy pregnant women. Therein, 393 cases were singletons and 47 were multiple pregnancies. Serum PSP levels were measured by specific enzyme-linked immunosorbent assay. The main outcome measures were serum PSP level (ng/mL) reference values in healthy pregnant women. RESULTS: The mean PSP reference values in women with singleton pregnancies were 7.9 ± 2.6 ng/mL (95% CI; 2.69-13.03 ng/mL). The PSP values in women with multiple pregnancies (9.17 ± 3.06 ng/mL (95% CI; 3.05-15.28 ng/mL)) were significantly higher (p = 0.001). The PSP values in the first trimester (6.94 ± 2.53 ng/mL) were lower compared to the second (7.42 ± 2.21 ng/mL) and third trimesters (8.33 ± 2.68 ng/mL, p = 0.0001). Subgroup analyses in singletons revealed no correlations between PSP values, maternal characteristics, and pre-existing medical conditions. CONCLUSION: The PSP values in healthy pregnant women (4-12 ng/mL) were in the range of the reference values of the general healthy population (8-16 ng/mL). This insight blazes a trail for further clinical studies on the use of PSP as a potential novel biomarker for the early detection of pregnancy-related diseases such as chorioamnionitis.

Identification of regenerating island-derived protein 3E in dogs.

Regenerating islet-derived protein (REG) 1A (aka pancreatic stone protein) and REG3A (aka pancreatitis-associated protein) are upregulated in humans with sepsis, pancreatitis, and gastrointestinal diseases, but little is known about this protein family in dogs. Our aim was to identify REG1 and REG3 family members in dogs. REG-family genes were computationally annotated in the canine genome and proteome, with verification of gene expression using publicly available RNA-seq data. The presence of the protein in canine pancreatic tissue and plasma was investigated with Western blot and immunohistochemistry, using anti-human REG1A and REG3A antibodies. Protein identity was confirmed with mass spectrometry. Two members of the REG3 subfamily were found in the canine genome, REG3E1 and REG3E2, both encoding for the same 176 AA protein, subsequently named REG3E. Anti-human REG3A antibodies demonstrated cross-reactivity with the canine REG3E protein in pancreas homogenates. In canine plasma, a protein band of approximately 17 kDa was apparent. Mass spectrometry confirmed this protein to be the product of the two annotated REG3E genes. Strong immunoreactivity to anti-human REG3A antibodies was found in sections of canine pancreas affected with acute pancreatitis, but it was weak in healthy pancreatic tissue. Recombinant canine REG3E protein underwent a selective trypsin digestion as described in other species. No evidence for the presence of a homolog of REG1A in dogs was found in any of the investigations. In conclusion, dogs express REG3E in the pancreas, whose role as biomarker merits further investigations. Homologs to human REG1A are not likely to exist in dogs.

Loss of LAT1 sex-dependently delays recovery after caerulein-induced acute pancreatitis.

BACKGROUND: The expression of amino acid transporters is known to vary during acute pancreatitis (AP) except for LAT1 (slc7a5), the expression of which remains stable. LAT1 supports cell growth by importing leucine and thereby stimulates mammalian target of rapamycin (mTOR) activity, a phenomenon often observed in cancer cells. The mechanisms by which LAT1 influences physiological and pathophysiological processes and affects disease progression in the pancreas are not yet known. AIM: To evaluate the role of LAT1 in the development of and recovery from AP. METHODS: AP was induced with caerulein (cae) injections in female and male mice expressing LAT1 or after its knockout (LAT1 Cre/LoxP). The development of the initial AP injury and its recovery were followed for seven days after cae injections by daily measuring body weight, assessing microscopical tissue architecture, mRNA and protein expression, protein synthesis, and enzyme activity levels, as well as by testing the recruitment of immune cells by FACS and ELISA. RESULTS: The initial injury, evaluated by measurements of plasma amylase, lipase, and trypsin activity, as well as the gene expression of dedifferentiation markers, did not differ between the groups. However, early metabolic adaptations that support regeneration at later stages were blunted in LAT1 knockout mice. Especially in females, we observed less mTOR reactivation and dysfunctional autophagy. The later regeneration phase was clearly delayed in female LAT1 knockout mice, which did not regain normal expression of the pancreas-specific differentiation markers recombining binding protein suppressor of hairless-like protein (rbpjl) and basic helix-loop-helix family member A15 (mist1). Amylase mRNA and protein levels remained lower, and, strikingly, female LAT1 knockout mice presented signs of fibrosis lasting until day seven. In contrast, pancreas morphology had returned to normal in wild-type littermates. CONCLUSION: LAT1 supports the regeneration of acinar cells after AP. Female mice lacking LAT1 exhibited more pronounced alterations than male mice, indicating a sexual dimorphism of amino acid metabolism.

De novo expression of gastrokines in pancreatic precursor lesions impede the development of pancreatic cancer.

Molecular events occurring in stepwise progression from pre-malignant lesions (pancreatic intraepithelial neoplasia; PanIN) to the development of pancreatic ductal adenocarcinoma (PDAC) are poorly understood. Thus, characterization of early PanIN lesions may reveal markers that can help in diagnosing PDAC at an early stage and allow understanding the pathology of the disease. We performed the molecular and histological assessment of patient-derived PanINs, tumor tissues and pancreas from mouse models with PDAC (KC mice that harbor K-RAS mutation in pancreatic tissue), where we noted marked upregulation of gastrokine (GKN) proteins. To further understand the role of gastrokine proteins in PDAC development, GKN-deficient KC mice were developed by intercrossing gastrokine-deficient mice with KC mice. Panc-02 (pancreatic cancer cells of mouse origin) were genetically modified to express GKN1 for further in vitro and in vivo analysis. Our results show that gastrokine proteins were absent in healthy pancreas and invasive cancer, while its expression was prominent in low-grade PanINs. We could detect these proteins in pancreatic juice and serum of KC mice. Furthermore, accelerated PanIN and tumor development were noted in gastrokine deficient KC mice. Loss of gastrokine 1 protein delayed apoptosis during carcinogenesis leading to the development of desmoplastic stroma while loss of gastrokine 2 increased the proliferation rate in precursor lesions. In summary, we identified gastrokine proteins in early pancreatic precursor lesions, where gastrokine proteins delay pancreatic carcinogenesis.

Analysis of Pain and Analgesia Protocols in Acute Cerulein-Induced Pancreatitis in Male C57BL/6 Mice.

Pancreatitis is known to be painful in humans and companion animals. However, the extent of pain in experimental mouse models of acute pancreatitis is unknown. Consequently, the severity classification of acute pancreatitis in mice is controversially discussed and standardized pain management is missing. In this study, we investigated acute Cerulein-induced pancreatitis with pain-specific and well-being orientated parameters to detect its impact on mice. Male C57BL/6J male mice were injected with Cerulein; animals that received saline injections served as control group. The animals were observed for weight change and water intake. To assess pain, behaviors like stretch-and-press and reduced rearing, the Mouse Grimace Scale, and von Frey hypersensitivity were assessed. Fecal corticosterone metabolites and burrowing behavior were assessed to detect changes in the animal's well-being. Pancreatitis severity was evaluated with amylase and lipase in the blood and pancreas histology. To investigate whether different analgesics can alleviate signs of pain, and if they influence pancreas inflammation, animals received Buprenorphine, Paracetamol in combination with Tramadol, or Metamizole in the drinking water. The calculated intake of these analgesics via drinking reached values stated to be efficient for pain alleviation. While pancreatitis did not seem to be painful, we detected acute pain from Cerulein injections that could not be alleviated by analgesics. The number of inflammatory cells in the pancreas did not differ with the analgesic administered. In conclusion: (1) Cerulein injections appear to be acutely painful but pain could not be alleviated by the tested analgesics, (2) acute pancreatitis induced by our protocol did not induce obvious signs of pain, (3) analgesic substances had no detectable influence on inflammation. Nevertheless, protocols inducing more severe or even chronic pancreatitis might evoke more pain and analgesic treatment might become imperative. Considering our results, we recommend the use of Buprenorphine via drinking water in these protocols. Further studies to search for efficient analgesics that can alleviate the acute pain induced by Cerulein injections are needed.

Attenuation of peripheral serotonin inhibits tumor growth and enhances immune checkpoint blockade therapy in murine tumor models.

Platelet-derived peripheral serotonin has pleiotropic effects on coagulation, metabolism, tissue regeneration, and cancer growth; however, the effect of serotonin on the tumor microenvironment remains understudied. Peripheral serotonin­deficient (Tph1−/−) mice displayed reduced growth of subcutaneous and orthotopically injected syngeneic murine pancreatic and colorectal cancers with enhanced accumulation of functional CD8+ T cells compared to control C57BL/6 mice, resulting in extended overall survival. Subcutaneous and orthotopic syngeneic tumors from Tph1−/− mice expressed less programmed cell death 1 ligand 1 (PD-L1), suggesting serotonin-mediated regulation. Serotonin enhanced expression of PD-L1 on mouse and human cancer cells in vitro via serotonylation, which is the formation of covalent bonds between glutamine residues and serotonin, resulting in activation of small G proteins. Serotonin concentrations in metastases of patients with abdominal tumors negatively correlated to the number of CD8+ tumor-infiltrating T cells. Depletion of serotonin cargo or inhibition of serotonin release from thrombocytes decreased growth of syngeneic pancreatic and colorectal tumors in wild-type mice, increased CD8+ T cell influx, and decreased PD-L1 expression. Pharmacological serotonin depletion with oral fluoxetine or intraperitoneal injection of the TPH1 inhibitor telotristat augmented the effects of programmed cell death protein 1 (PD-1) checkpoint blockade and triggered long-term tumor control in mice subcutaneously inoculated with syngeneic colorectal and pancreatic tumors. Overall, peripheral serotonin weakens effector functions of CD8+ T cells within tumors. Clinically approved serotonin targeting agents alone or in combination with PD-1 blockade provided long-term control of established tumors in murine models, warranting further investigation of the clinical translatability of these findings.

Single or combined ablation of peripheral serotonin and p21 limit adipose tissue expansion and metabolic alterations in early adulthood in mice fed a normocaloric diet.

Identifying the fundamental molecular factors that drive weight gain even in the absence of hypercaloric food intake, is crucial to enable development of novel treatments for the global pandemic of obesity. Here we investigated both adipose tissue-specific and systemic events that underlie the physiological weight gain occurring during early adulthood in mice fed a normocaloric diet. In addition, we used three different genetic models to identify molecular factors that promote physiological weight gain during normocaloric and hypercaloric diets. We demonstrated that normal physiological weight gain was accompanied by an increase in adipose tissue mass and the presence of cellular and metabolic signatures typically found during obesity, including adipocyte hypertrophy, macrophage recruitment into visceral fat and perturbed glucose metabolism. At the molecular level, this was associated with an increase in adipose tissue tryptophan hydroxylase 1 (Tph1) transcripts, the key enzyme responsible for the synthesis of peripheral serotonin. Genetic inactivation of Tph1 was sufficient to limit adipose tissue expansion and associated metabolic alterations. Mechanistically, we discovered that Tph1 inactivation resulted in down-regulation of cyclin-dependent kinase inhibitor p21Waf1/Cip1 expression. Single or double ablation of Tph1 and p21 were equally effective in preventing adipocyte expansion and systemic perturbation of glucose metabolism, upon both normocaloric and hypercaloric diets. Our results suggest that serotonin and p21 act as a central molecular determinant of weight gain and associated metabolic alterations, and highlights the potential of targeting these molecules as a pharmacologic approach to prevent the development of obesity.

Incidence and Time Point of Sepsis Detection as Related to Different Sepsis Definitions in Severely Burned Patients and Their Accompanying Time Course of Pro-Inflammatory Biomarkers.

BACKGROUND: Diagnosis of sepsis in burn patients remains difficult for various reasons. One major problem is the definition of sepsis itself. Therefore, previous and current sepsis definitions are a matter of ongoing validation, but a well-defined consensus on which clinical and laboratory parameters to incorporate in such a definition is lacking. The aim of the present study was to compare the incidence and time-related occurrence of septic events according to different definitions as well as their accompanying time course of pro-inflammatory biomarkers. METHODS: Across the first 14 days after admission, the incidence and time point of sepsis according to three different definitions (Sepsis-3, Sepsis American Burns Association [ABA] 2007, Sepsis Zurich Burn Center) were assessed on a daily basis in adult burn patients with total body surface area (TBSA) ≥15% admitted to the Zurich Burn Center between May 2015 and October 2018. In order to investigate how well daily drawn proinflammatory biomarkers (white blood cells (WBCs), C-reactive protein (CRP), procalcitonin (PCT), and novel pancreatic stone protein (PSP)) reflect the progression of sepsis depending on its type of definition, a longitudinal mixed model analysis was performed across the first 14 days for septic and non-septic patients. Additionally, the relative increase of biomarker levels 24, 48, and 72 h prior to a septic event was analyzed for each definition used. RESULTS: In our cohort of 90 severely burned patients, Sepsis-3 identified 46 patients (51.1%) as septic, while ABA 2007 and the Zurich Burn Center definition counted 33 patients (36.7%) and 24 patients (26.6%), respectively. Sepsis-3 detected sepsis about 1 day earlier than Sepsis ABA 2007 (p < 0.001) and about 0.5 days earlier than Sepsis Zurich Burn Center (p = 0.04). The course of pro-inflammatory biomarkers was largely unaffected by the type of sepsis definition. Irrespective of the sepsis definition, PSP was the only marker to demonstrate a highly significant interaction between time and group (sepsis versus no sepsis) (p < 0.001) with a 3.3-5.5-fold increase within 72 h before the event of sepsis, whereas CRP, PCT, and WBC showed only mild undulations. CONCLUSIONS: Despite the ongoing dilemma of how to define sepsis in burn patients, a continually calculated SOFA score as used in Sepsis-3 is advantageous to early identify a patient's detrimental progression to sepsis. Inclusion of biomarkers, such as PSP, may help support the burn specialist's diagnosis of sepsis and could improve the diagnostic performance of current and future definitions in burn patients.

Phase Ib dose-escalation study of the hypoxia-modifier Myo-inositol trispyrophosphate in patients with hepatopancreatobiliary tumors.

Hypoxia is prominent in solid tumors and a recognized driver of malignancy. Thus far, targeting tumor hypoxia has remained unsuccessful. Myo-inositol trispyrophosphate (ITPP) is a re-oxygenating compound without apparent toxicity. In preclinical models, ITPP potentiates the efficacy of subsequent chemotherapy through vascular normalization. Here, we report the results of an unrandomized, open-labeled, 3 + 3 dose-escalation phase Ib study (NCT02528526) including 28 patients with advanced primary hepatopancreatobiliary malignancies and liver metastases of colorectal cancer receiving nine 8h-infusions of ITPP over three weeks across eight dose levels (1'866-14'500 mg/m2/dose), followed by standard chemotherapy. Primary objectives are assessment of the safety and tolerability and establishment of the maximum tolerated dose, while secondary objectives include assessment of pharmacokinetics, antitumor activity via radiological evaluation and assessment of circulatory tumor-specific and angiogenic markers. The maximum tolerated dose is 12,390 mg/m2, and ITPP treatment results in 32 treatment-related toxicities (mostly hypercalcemia) that require little or no intervention. 52% of patients have morphological disease stabilization under ITPP monotherapy. Following subsequent chemotherapy, 10% show partial responses while 60% have stable disease. Decreases in angiogenic markers are noted in ∼60% of patients after ITPP and tend to correlate with responses and survival after chemotherapy.

Accuracy of pancreatic stone protein for the diagnosis of infection in hospitalized adults: a systematic review and individual patient level meta-analysis.

BACKGROUND: Accurate biomarkers to diagnose infection are lacking. Studies reported good performance of pancreatic stone protein (PSP) to detect infection. The objective of the study was to determine the performance of PSP in diagnosing infection across hospitalized patients and calculate a threshold value for that purpose. METHODS: A systematic search across Cochrane Central Register of Controlled Trials and MEDLINE databases (1966-March 2019) for studies on PSP published in English using 'pancreatic stone protein', 'PSP', 'regenerative protein', 'lithostatin' combined with 'infection' and 'sepsis' found 44 records. The search was restricted to the five trials that evaluated PSP for the initial detection of infection in hospitalized adults. Individual patient data were obtained from the investigators of all eligible trials. Data quality and validity was assessed according to PRISMA guidelines. We choose a fixed-effect model to calculate the PSP cut-off value that best discriminates infected from non-infected patients. RESULTS: Infection was confirmed in 371 of 631 patients. The median (IQR) PSP value of infected versus uninfected patients was 81.5 (30.0-237.5) versus 19.2 (12.6-33.57) ng/ml, compared to 150 (82.70-229.55) versus 58.25 (15.85-120) mg/l for C-reactive protein (CRP) and 0.9 (0.29-4.4) versus 0.15 (0.08-0.5) ng/ml for procalcitonin (PCT). Using a PSP cut-off of 44.18 ng/ml, the ROC AUC to detect infection was 0.81 (0.78-0.85) with a sensitivity of 0.66 (0.61-0.71), specificity of 0.83 (0.78-0.88), PPV of 0.85 (0.81-0.89) and NPV of 0.63 (0.58-0.68). When a model combining PSP and CRP was used, the ROC AUC improved to 0.90 (0.87-0.92) with higher sensitivity 0.81 (0.77-0.85) and specificity 0.84 (0.79-0.90) for discriminating infection from non-infection. Adding PCT did not improve the performance further. CONCLUSIONS: PSP is a promising biomarker to diagnose infections in hospitalized patients. Using a cut-off value of 44.18 ng/ml, PSP performs better than CRP or PCT across the considered studies. The combination of PSP with CRP further enhances its accuracy.

Tumor Oxygenation by Myo-Inositol Trispyrophosphate Enhances Radiation Response.

PURPOSE: Tumor hypoxia is a major limiting factor for successful radiation therapy outcomes, with hypoxic cells being up to 3-fold more radiation resistant than normoxic cells; tumor hypoxia creates a tumor microenvironment that is hostile to immune response. Thus, pharmaceutical-induced tumor oxygenation before radiation therapy represents an interesting method to enhance the efficacy of radiation therapy. Myo-inositol trispyrophosphate (ITPP) triggers a decrease in the affinity of oxygen to hemoglobin, which leads to an increased release of oxygen upon tissue demand, including in hypoxic tumors. METHODS AND MATERIALS: The combined treatment modality of high-dose bolus ITPP with a single high-dose fraction of ionizing radiation (IR) was investigated for its mechanics and efficacy in multiple preclinical animal tumor models in immunocompromised and immunocompetent mice. The dynamics of tumor oxygenation were determined by serial hypoxia-oriented bioimaging. Initial and residual DNA damage and the integrity of the tumor vasculature were quantified on the immunohistochemical level in response to the different treatment combinations. RESULTS: ITPP application did not affect tumor growth as a single treatment modality, but it rapidly induced tumor oxygenation, as demonstrated by in vivo imaging, and significantly reduced tumor growth when combined with IR. An immunohistochemical analysis of γH2AX foci demonstrated increased initial and residual IR-induced DNA damage as the primary mechanism for radiosensitization within initially hypoxic but ITPP-oxygenated tumor regions. Scheduling experiments revealed that ITPP increases the efficacy of ionizing radiation only when applied before radiation therapy. Irradiation alone damaged the tumor vasculature and increased tumor hypoxia, which were both prevented by combined treatment with ITPP. Interestingly, the combined treatment modality also promoted increased immune cell infiltration. CONCLUSIONS: ITPP-mediated tumor oxygenation and vascular protection triggers immediate and delayed processes to enhance the efficacy of ionizing radiation for successful radiation therapy.

Pancreatic Stone Protein Predicts Sepsis in Severely Burned Patients Irrespective of Trauma Severity: A Monocentric Observational Study.

OBJECTIVE: The burn victim's inherent state of hyperinflammation frequently camouflages septic events delaying the initiation of targeted intensive care therapy. Accurate biomarkers are urgently needed to support sepsis detection before patients' clinical deterioration. SUMMARY OF BACKGROUND DATA: Evidence on the usefulness of pancreatic stone protein (PSP) as a powerful diagnostic and prognostic marker in critically ill patients has recently accumulated. METHODS: Analysis of biomarker kinetics (PSP, routine markers) was performed on 90 patients admitted to the Zurich Burn Center between May 2015 and October 2018 with burns ≥15% total body surface area with regard to infection and sepsis (Sepsis-3) over a 14-day time course. RESULTS: PSP differentiated between sepsis, infection and sterile inflammation from day 3 onward with an area under the curve of up to 0.89 (P < 0.001), therefore, competing with procalcitonin (area under the curve = 0.86, P < 0.001). Compared to routine inflammatory biomarkers, only PSP demonstrated a significant interaction between time and presence of sepsis - signifying a steeper increase in PSP levels in septic patients as opposed to those exhibiting a nonseptic course (interaction P < 0.001). Event-related analysis demonstrated tripled PSP serum levels within 72 hours and doubled levels within 48 hours before a clinically apparent sepsis. CONCLUSION: PSP is able to differentiate between septic and nonseptic patients during acute burn care. Its steep rise up to 72 hours before clinically overt deterioration has the potential for physicians to timely initiate treatment with reduced mortality and costs.

Response of routine inflammatory biomarkers and novel Pancreatic Stone Protein to inhalation injury and its interference with sepsis detection in severely burned patients.

BACKGROUND: Inhalation of thermal and chemical products of combustion evokes an immune response measurable at a systemic level. Inhalation injury related kinetics of currently available inflammatory biomarkers and novel Pancreatic Stone Protein (PSP) as well as their interference with septic events has not been addressed to literature yet. METHODS: Analysis of the influence of inhalation injury and ARDS on biomarker kinetics (PSP, procalcitonin (PCT), C-reactive Protein (CRP), white blood cells (WBC)) in 90 patients admitted to Zurich Burn Center between May 2015 and October 2018 with burns ≥15% total body surface area (TBSA) over 14 days. RESULTS: Twenty-five (27%) of 90 included patients presented with inhalation injury (median age 52 years [IQR 27], median TBSA 31.5% [IQR 21], mean ABSI-Score 7±3). At admission, only WBC demonstrated significantly higher values in the inhalation injury group (p=0.011). Acute respiratory distress syndrome (ARDS) was present in 32% without association to the severity of inhalation injury (p=0.11). WBC, CRP and PCT failed to delineate inhalation injury related inflammation from septic progression at most time points. PSP was the strongest marker to identify septic patients both by its higher values and steeper increase over time (p<0.001). CONCLUSION: Inhalation injury leads to an inflammatory response at a systemic level with alterations of biomarkers. While routine inflammatory markers demonstrated strong interferences between inhalation injury with its associated ARDS and evolving sepsis, PSP reliably identified septic patients in a setting of inflammatory turbulences secondary to inhalation injury.

Amelioration of Murine Autoimmune Pancreatitis by Targeted LTßR Inhibition and Anti-CD20 Treatment.

Autoimmune pancreatitis (AIP) is a rare form of chronic pancreatitis, for which treatment options, especially the long-term management, are limited. The only therapy that has been established and accepted so far is corticosteroids, but the relapse rate is significant. In the current study, we discern the effector mechanisms of targeted LTßR pathway inhibition using LTßR-Ig. Furthermore, the efficacy of LTßR-Ig therapy is compared with the depletion of immune cell subsets (CD4+ and CD20+), which are suggested to play a pathological role in AIP development. Three well-established mouse models of AIP were used to examine treatment efficacies and mechanisms. Tg(Ela1-Lta,b) mice represent a genetic model, in which AIP develops spontaneously. In MRL/Mp and IL-10-/- mice, AIP is induced by repeated polyinosinic:polycytidylic acid injection. Mice with AIP were treated with anti-CD20, anti-CD4 mAbs, or targeted LTßR-Ig. LTßR-Ig and anti-CD20 treatment led to significant improvement of AIP, including a decrease in autoantibody production and pancreatic inflammation in Tg(Ela1-Lta,b) and IL-10-/- mice. The molecular mechanism of this beneficial effect possibly involves the downregulation of Stat3 and noncanonical NF-κb activation. Anti-CD4 treatment reduced Th1 and Th2 signature but did not alleviate AIP. Additionally, in contrast to anti-CD20 or anti-CD4 treatments, blocking LTßR signaling disrupted tertiary lymphoid organs in all three models. We demonstrate that treatment with LTßR-Ig or anti-CD20 Ab alleviated murine AIP. LTßR-Ig treatment for AIP was effective in both lymphotoxin-dependent and lymphotoxin-independent AIP models, possibly because of its dual anti-inflammatory and antiautoimmune mechanisms.

Large-Scale Profiling of Signaling Pathways Reveals a Distinct Demarcation between Normal and Extended Liver Resection.

Despite numerous studies addressing normal liver regeneration, we still lack comprehensive understanding of the biological processes underlying failed liver regeneration. Therefore, we analyzed the activity of 271 intracellular signaling pathways (ISPs) by genome wide profiling of differentially expressed RNAs in murine liver tissue biopsies after normal hepatectomy (nHx; 68% of liver removed) and extended hepatectomy (eHx; 86% of liver removed). Comprehensive, genome-wide transcriptome profiling using RNAseq was performed in liver tissue obtained from mice (sham, nHx, and eHx) harvested 1, 8, 16, 32, and 48 h after operation (n = 3 per group) and the OncoFinder toolkit was used for an unsupervised, unbiased identification of intracellular signaling pathways (ISP) activity. We observed that the normal regenerative process requires a transient activation and silencing of approximately two dozen of ISPs. After nHx, the Akt Pathway represented with 13 branches, the Chromatin Pathway and the DDR Pathways dominated. After eHx, the ATM main pathway and two of its branches (Cell Survival; G2_M Checkpoint Arrest) dominated, as well as the Hypoxia Pathways. Further, 14 ISPs demonstrated a strong inverse regulation, with the Hedgehog and the Brca1 Main Pathways as chief activators after nHx, and the ATM Pathway (G2_M Checkpoint Arrest) as the dominating constraining response after eHx.

Serine administration as a novel prophylactic approach to reduce the severity of acute pancreatitis during diabetes in mice.

AIMS/HYPOTHESIS: Compared with the general population, individuals with diabetes have a higher risk of developing severe acute pancreatitis, a highly debilitating and potentially lethal inflammation of the exocrine pancreas. In this study, we investigated whether 1-deoxysphingolipids, atypical lipids that increase in the circulation following the development of diabetes, exacerbate the severity of pancreatitis in a diabetic setting. METHODS: We analysed whether administration of an L-serine-enriched diet to mouse models of diabetes, an established method for decreasing the synthesis of 1-deoxysphingolipids in vivo, reduced the severity of acute pancreatitis. Furthermore, we elucidated the molecular mechanisms underlying the lipotoxicity exerted by 1-deoxysphingolipids towards rodent pancreatic acinar cells in vitro. RESULTS: We demonstrated that L-serine supplementation reduced the damage of acinar tissue resulting from the induction of pancreatitis in diabetic mice (average histological damage score: 1.5 in L-serine-treated mice vs 2.7 in the control group). At the cellular level, we showed that L-serine decreased the production of reactive oxygen species, endoplasmic reticulum stress and cellular apoptosis in acinar tissue. Importantly, these parameters, together with DNA damage, were triggered in acinar cells upon treatment with 1-deoxysphingolipids in vitro, suggesting that these lipids are cytotoxic towards pancreatic acinar cells in a cell-autonomous manner. In search of the initiating events of the observed cytotoxicity, we discovered that 1-deoxysphingolipids induced early mitochondrial dysfunction in acinar cells, characterised by ultrastructural alterations, impaired oxygen consumption rate and reduced ATP synthesis. CONCLUSIONS/INTERPRETATION: Our results suggest that 1-deoxysphingolipids directly damage the functionality of pancreatic acinar cells and highlight that an L-serine-enriched diet may be used as a promising prophylactic intervention to reduce the severity of pancreatitis in the context of diabetes.