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Background: Hepatic venous pressure gradient (HVPG) is the gold standard index for evaluating portal hypertension; however, measuring HVPG is invasive. Although transient elastography (TE) is the most common procedure for evaluating organ stiffness, accurate measurement of spleen stiffness (SS) is difficult. We developed a device to demonstrate the diagnostic precision of TE and suggest this technique as a valuable new method to measure SS., Methods: Of 292 consecutive patients enrolled in this single-centre, translational, cross-sectional study from June through September in 2019, 200 underwent SS measurement (SSM) using an M probe (training set, n = 130; inspection set, n = 70). We performed TE with B-mode imaging using an ultrasound-fusion method, printed new devices with a three-dimensional printer, and attached the magnetic position sensor to the convex and M probes. We evaluated the diagnostic precision of TE to evaluate the risk of esophagogastric varices (EGVs)., Results: The median spleen volume was 245 mL (range, 64-1,720 mL), and it took 2 minutes to acquire a B-mode image using the ultrasound-fusion method. The median success rates of TE were 83.3% and 57.6% in patients with and without the new device, respectively (p<0.001); it was 76.9% and 35.0% in patients with and without splenomegaly (<100 mL), respectively (p<0.001). In the prediction of EGVs, the areas under the receiver operating characteristic curve were 0.921 and 0.858 in patients with and without the new device, respectively (p = 0.043). When the new device was attached, the positive and negative likelihood ratios were 3.44 and 0.11, respectively. The cut-off value of SSM was 46.0 kPa. Data that were similar between the validation and training sets were obtained., Conclusions: The SS can be precisely measured using this new device with TE and ultrasound-fusion method. Similarly, we can estimate the bleeding risk due to EGV using this method., Competing Interests: The authors have declared that no competing interests exist.

Purpose: Haematological toxicities occur in patients receiving oxaliplatin. Mild anaemia (grade 1-2) is a common side effect and approximately 90% of recipients develop measurable spleen enlargement. Although generally asymptomatic, oxaliplatin-induced splenomegaly is independently associated with complications following liver resection for colorectal liver metastasis and separately with poorer patient outcomes. Here, we investigated oxaliplatin-induced haematological toxicities and splenomegaly in mice treated with escalating dosages comparable to those prescribed to colorectal cancer patients., Methods: Blood was analysed, and smears assessed using Wright-Giemsa staining. Paw coloration was quantified as a marker of anaemia. Spleen weight and morphology were assessed for abnormalities relating to splenomegaly and a flow cytometry and multiplex cytokine array assessment was performed on splenocytes. The liver was assessed for sinusoidal obstructive syndrome., Results: Blood analysis showed dose dependent decreases in white and red blood cell counts, and significant changes in haematological indices. Front and hind paws exhibited dose dependent and dramatic discoloration indicative of anaemia. Spleen weight was significantly increased indicating splenomegaly, and red pulp tissue exhibited substantial dysplasia. Cytokines and chemokines within the spleen were significantly affected with temporal upregulation of IL-6, IL-1α and G-CSF and downregulation of IL-1β, IL-12p40, MIP-1β, IL-2 and RANTES. Flow cytometric analysis demonstrated alterations in splenocyte populations, including a significant reduction in CD45+ cells. Histological staining of the liver showed no evidence of sinusoidal obstructive syndrome but there were signs suggestive of extramedullary haematopoiesis., Conclusion: Chronic oxaliplatin treatment dose dependently induced haematological toxicity and splenomegaly characterised by numerous physiological and morphological changes, which occurred independently of sinusoidal obstructive syndrome., Competing Interests: The authors have declared that no competing interests exist.

To confirm that neoplastic monocyte-derived collagen- and fibronectin-producing fibrocytes induce bone marrow (BM) fibrosis in primary myelofibrosis (PMF), we injected PMF BM-derived fibrocyte-precursor CD14+/CD34- monocytes into the tail vein of NOD-SCID-γ (NSG) mice. PMF BM-derived CD14+/CD34- monocytes engrafted and induced a PMF-like phenotype with splenomegaly, myeloid hyperplasia with clusters of atypical megakaryocytes, persistence of the JAK2V617F mutation, and BM and spleen fibrosis. As control we used normal human BM-derived CD14+/CD34- monocytes. These monocytes also engrafted and gave rise to normal megakaryocytes that, like PMF CD14+/CD34--derived megakaryocytes, expressed HLA-ABC and human CD42b antigens. Using 2 clonogenic assays we confirmed that PMF and normal BM-derived CD14+/CD34- monocytes give rise to megakaryocyte colony-forming cells, suggesting that a subpopulation BM monocytes harbors megakaryocyte progenitor capacity. Taken together, our data suggest that PMF monocytes induce myelofibrosis-like phenotype in immunodeficient mice and that PMF and normal BM-derived CD14+/CD34- monocytes give rise to megakaryocyte progenitor cells., Competing Interests: The authors have declared that no competing interests exist.

GPR116 (ADGRF5) and ELTD1 (ADGRL4) belong to different subfamilies of the adhesion G-protein-coupled receptor group but are both expressed in endothelial cells. We therefore analyzed their functions in mice lacking these receptors. While loss of GPR116 or ELTD1 alone had no obvious effect on cardiovascular or kidney function, mice lacking both, GPR116 and ELTD1, showed malformations of the aortic arch arteries and the cardiac outflow tract leading to perinatal lethality in about 50% of the mutants. In addition to cardiovascular malformations, surviving mice developed renal thrombotic microangiopathy as well as hemolysis and splenomegaly, and their lifespan was significantly reduced. Loss of GPR116 and ELTD1 specifically in endothelial cells or neural crest-derived cells did not recapitulate any of the phenotypes observed in GPR116-ELTD1 double deficient mice, indicating that loss of GPR116 and ELTD1 expressed by other cells accounts for the observed cardiovascular and renal defects.

Background: Spleen enlargement is often detected in patients with liver cirrhosis, but the precise pathogenetic mechanisms behind the phenomenon have not been clearly elucidated. We investigated the pathogenetic mechanisms of splenomegaly in both portal hypertensive patients and rats, and tried to identify the possible therapy for this disease., Methods: Spleen samples were collected from portal hypertensive patients after splenectomy. Rat models of portal hypertension were induced by common bile duct ligation and partial portal vein ligation. Spleen samples from patients and rats were used to study the characteristics of splenomegaly by histological, immunohistochemical, and western blot analyses. Rapamycin or vehicle was administered to rats to determine the contribution of mTOR signaling pathway in the development of splenomegaly., Results: We found that not only spleen congestion, but also increasing angiogenesis, fibrogenesis, inflammation and proliferation of splenic lymphoid tissue contributed to the development of splenomegaly in portal hypertensive patients and rats. Intriguingly, splenomegaly developed time-dependently in portal hypertensive rat that accompanied with progressive activation of mTOR signaling pathway. mTOR blockade by rapamycin profoundly ameliorated splenomegaly by limiting lymphocytes proliferation, angiogenesis, fibrogenesis and inflammation as well as decreasing portal pressure., Conclusions: This study provides compelling evidence indicating that mTOR signaling activation pathway plays a key role in the pathogenesis of splenomegaly in both portal hypertensive patients and rats. Therapeutic intervention targeting mTOR could be a promising strategy for patients with portal hypertension and splenomegaly.

Chromosomal translocations are driver mutations of human cancers, particularly leukemias. They define disease subtypes and are used as prognostic markers, for minimal residual disease monitoring and therapeutic targets. Due to their low incidence, several translocations and their biological consequences remain poorly characterized. To address this, we engineered mouse strains that conditionally express E2A-HLF, a fusion oncogene from the translocation t(17;19) associated with 1% of pediatric B-cell precursor ALL. Conditional oncogene activation and expression were directed to the B-cell compartment by the Cre driver promoters CD19 or Mb1 (Igα, CD79a), or to the hematopoietic stem cell compartment by the Mx1 promoter. E2A-HLF expression in B-cell progenitors induced hyposplenia and lymphopenia, whereas expression in hematopoietic stem/progenitor cells was embryonic lethal. Increased cell death was detected in E2A-HLF expressing cells, suggesting the need for cooperating genetic events that suppress cell death for B-cell oncogenic transformation. E2A-HLF/Mb1.Cre aged mice developed a fatal myeloproliferative-like disorder with low frequency characterized by leukocytosis, anemia, hepatosplenomegaly and organ-infiltration by mature myelocytes. In conclusion, we have developed conditional E2A-HLF knock-in mice, which provide an experimental platform to study cooperating genetic events and further elucidate translational biology in cross-species comparative studies.

Oxaliplatin-based chemotherapy is widely used to treat advanced colorectal cancer (CRC). Sinusoidal obstruction syndrome (SOS) due to oxaliplatin is a serious type of chemotherapy-associated liver injury (CALI) in CRC patients. SOS is thought to be caused by the sinusoidal endothelial cell damage, which results in the release of unusually-large von Willebrand factor multimers (UL-VWFMs) from endothelial cells. To investigate the pathophysiology of CALI after oxaliplatin-based chemotherapy, we analyzed plasma concentration of von Willebrand factor (VWF) and the distribution of VWFMs in CRC patients. Twenty-three patients with advanced CRC who received oxaliplatin-based chemotherapy with (n = 6) and without (n = 17) bevacizumab were analyzed. CALI (n = 6) and splenomegaly (n = 9) were found only in patients who did not treated with bevacizumab. Plasma VWF antigen (VWF:Ag) and serum aspartate aminotransferase (AST) levels increased after chemotherapy only in patients without bevacizumab. VWFM analysis in patients who did not receive bevacizumab showed the presence of UL-VWFMs and absence of high molecular weight VWFMs during chemotherapy, especially in those with CALI. In addition, plasma VWF:Ag and AST levels increased after chemotherapy in patients with splenomegaly (n = 9), but not in patients without splenomegaly (n = 14). Histological findings in the liver tissue of patients who did not receive bevacizumab included sinusoidal dilatation and microthrombi in the sinusoids. Many microthrombi were positive for both anti-IIb/IIIa and anti-VWF antibodies. Plasma UL-VWFM levels might be increased by damage to endothelial cells as a result of oxaliplatin-based chemotherapy. Bevacizumab could prevent CALI and splenomegaly through inhibition of VWF-rich platelet thrombus formation.

Splenomegaly and pancytopenia are common in Wilson's disease (WD) and splenectomy is one of the conventional treatments for splenomegaly and the associated pancytopenia. However, splenectomy remained controversial for hypersplenism in WD as it was reported that splenectomy leaded to serious emotional and neurological deterioration in WD patients with hypersplenism. In the current study, we present our experiences in 70 WD patients with hypersplenism who had undergone splenectomy, outlining the safety and efficacy of splenectomy in WD. The clinical database of 70 WD patients with hypersplenism who had undergone splenectomy in our hospital between 2009 and 2013 were reviewed and followed-up regularly. Before splenectomy, all the patients accepted a short period of anti-copper treatment with intravenous sodium 2, 3-dimercapto-1-propane sulfonate (DMPS). All the patients demonstrated a marked improvement in platelet and leucocyte counts after splenectomy. No severe postoperative complication was observed. In particular, none of the 37 patients with mixed neurologic and hepatic presentations experienced neurological deterioration after splenectomy, and none of the patients with only hepatic presentations newly developed neurological symptoms. During the one year follow-up period, no patient presented hepatic failure or hepatic encephalopathy, no hepatic patient newly developed neurological presentations, and only 3 patients with mixed neurologic and hepatic presentations suffered neurological deterioration and these 3 patients had poor compliance of anti-copper treatment. Quantative analysis of the neurological symptoms in the 37 patients using the Unified Wilson's Disease Rating Scale (UWDRS) showed that the neurological symptoms were not changed in a short-term of one week after splenectomy but significantly improved in a long-term of one year after splenectomy. Additionally, compared to that before splenectomy, the esophageal gastric varices in most patients significantly improved one year after splenectomy. Thus, we may conclude that splenectomy is a safe and effective therapeutic measure for hypersplenism in WD patients who had been preoperatively treated with DMPS for powerful anti-copper therapy.

Differentiation of infectious causes in severely ill children is essential but challenging in sub- Saharan Africa. The aim of the study was to determine clinical indicators that are able to identify bacterial co-infections in P. falciparum infected children in rural Ghana. In total, 1,915 severely ill children below the age of 15 years were recruited at Agogo Presbyterian Hospital in Ghana between May 2007 and February 2011. In 771 (40%) of the children malaria parasites were detected. This group was analyzed for indicators of bacterial co-infections using bivariate and multivariate regression analyses with 24 socio-economic variables, 16 terms describing medical history and anthropometrical information and 68 variables describing clinical symptoms. The variables were tested for sensitivity, specificity, positive predictive value and negative predictive value. In 46 (6.0%) of the children with malaria infection, bacterial co-infection was detected. The most frequent pathogens were non-typhoid salmonellae (45.7%), followed by Streptococcus spp. (13.0%). Coughing, dehydration, splenomegaly, severe anemia and leukocytosis were positively associated with bacteremia. Domestic hygiene and exclusive breastfeeding is negatively associated with bacteremia. In cases of high parasitemia (>10,000/μl), a significant association with bacteremia was found for splenomegaly (OR 8.8; CI 1.6-48.9), dehydration (OR 18.2; CI 2.0-166.0) and coughing (OR 9.0; CI 0.7-118.6). In children with low parasitemia, associations with bacteremia were found for vomiting (OR 4.7; CI 1.4-15.8), severe anemia (OR 3.3; CI 1.0-11.1) and leukocytosis (OR 6.8 CI 1.9-24.2). Clinical signs of impaired microcirculation were negatively associated with bacteremia. Ceftriaxone achieved best coverage of isolated pathogens. The results demonstrate the limitation of clinical symptoms to determine bacterial co-infections in P. falciparum infected children. Best clinical indicators are dependent on the parasitemia level. Even with a moderate sensitivity of >60%, only low positive predictive values can be obtained due to low prevalence of bacteremia. Rapid testing for distinguishing parasitemia and bacteremia is essential.

Background: The aim of this study was to evaluate the accuracy, safety, and role of splenic biopsy in the management of patients with non-mass-forming isolated splenomegaly and suspected malignant lymphoma., Methods: Between 2001 and 2013, 137 biopsies were performed under computed tomography (CT) fluoroscopic guidance in 39 patients. All patients had splenomegaly based on the CT findings and a suspected diagnosis of malignant lymphoma based on their clinical symptoms. The spleen was the only accessible site to perform a biopsy, and no mass lesions could be identified in the spleen., Results: The overall sensitivity, specificity, and diagnostic accuracy of image-guided biopsy for malignant lymphoma were 88%, 100% and 92%, respectively. Major complications occurred in 3 patients. In 1 patient, transcatheter arterial embolization was performed due to hemorrhage, and two patients needed blood transfusion because of hematoma development, without the need for further treatment., Conclusions: Image-guided splenic core-needle biopsy is a safe and accurate technique with a high diagnostic accuracy in most patients who with non-mass-forming isolated splenomegaly and suspected underlying malignant lymphoma.

Objective: Hepatic stellate cells (HSCs) transdifferentiation and subsequent inflammation are important pathological processes involved in the formation of cirrhotic portal hypertension. This study characterizes the pathogenetic mechanisms leading to cholestatic liver fibrosis and portal hypertension, and focuses on mammalian target of rapamycin (mTOR) pathway as a potential modulator in the early phase of cirrhotic portal hypertension., Methods: Early cirrhotic portal hypertension was induced by bile duct ligation (BDL) for three weeks. One week after operation, sham-operated (SHAM) and BDL rats received rapamycin (2 mg/kg/day) by intraperitoneal injection for fourteen days. Vehicle-treated SHAM and BDL rats served as controls. Fibrosis, inflammation, and portal pressure were evaluated by histology, morphometry, and hemodynamics. Expressions of pro-fibrogenic and pro-inflammatory genes in liver were measured by RT-PCR; alpha smooth muscle actin (α-SMA) and antigen Ki67 were detected by immunohistochemistry; expressions of AKT/mTOR signaling molecules, extracellular-signal-regulated kinase 1/2 (ERK1/2), p-ERK1/2, and interleukin-1 beta (IL-1β) were assessed by western blot., Results: The AKT/mTOR signaling pathway was markedly activated in the early phase of cirrhotic portal hypertension induced by BDL in rats. mTOR blockade by rapamycin profoundly improved liver function by limiting inflammation, fibrosis and portal pressure. Rapamycin significantly inhibited the expressions of phosphorylated 70KD ribosomal protein S6 kinase (p-P70S6K) and phosphorylated ribosomal protein S6 (p-S6) but not p-AKT Ser473 relative to their total proteins in BDL-Ra rats. Those results suggested that mTOR Complex 1 (mTORC1) rather than mTORC2 was inhibited by rapamycin. Interestingly, we also found that the level of p-ERK1/2 to ERK1/2 was significantly increased in BDL rats, which was little affected by rapamycin., Conclusions: The AKT/mTOR signaling pathway played an important role in the early phase of cirrhotic portal hypertension in rats, which could be a potential target for therapeutic intervention in the early phase of such pathophysiological progress.

Background: Pancreatic cancer is one of the most aggressive cancers, with tumor-induced myeloid-derived suppressor cells (MDSC) contributing to its pathogenesis and ineffective therapies. In response to cytokine/chemokine receptor activation, src homology 2 domain-containing inositol 5'-phosphatase-1 (SHIP-1) influences phosphatidylinositol-3-kinase (PI3K) signaling events, which regulate immunohomeostasis. We hypothesize that factors from murine pancreatic cancer cells cause the down-regulation of SHIP-1 expression, which may potentially contribute to MDSC expansion, and the suppression of CD8(+) T cell immune responses. Therefore, we sought to determine the role of SHIP-1 in solid tumor progression, such as murine pancreatic cancer., Methodology and Principal Findings: Immunocompetent C57BL/6 mice were inoculated with either murine Panc02 cells (tumor-bearing [TB] mice) or Phosphate Buffer Saline (PBS) (control mice). Cytometric Bead Array (CBA) analysis of supernatants of cultured Panc02 detected pro-inflammatory cytokines such as IL-6, IL-10 and MCP-1. TB mice showed a significant increase in serum levels of pro-inflammatory factors IL-6 and MCP-1 measured by CBA. qRT-PCR and Western blot analyses revealed the in vivo down-regulation of SHIP-1 expression in splenocytes from TB mice. Western blot analyses also detected reduced SHIP-1 activity, increased AKT-1 and BAD hyper-phosphorylation and up-regulation of BCL-2 expression in splenocytes from TB mice. In vitro, qRT-PCR and Western blot analyses detected reduced SHIP-1 mRNA and protein expression in control splenocytes co-cultured with Panc02 cells. Flow cytometry results showed significant expansion of MDSC in peripheral blood and splenocytes from TB mice. AutoMACS sorted TB MDSC exhibited hyper-phosphorylation of AKT-1 and over-expression of BCL-2 detected by western blot analysis. TB MDSC significantly suppressed antigen-specific CD8(+) T cell immune responses in vitro., Conclusion/significance: SHIP-1 may regulate immune development that impacts MDSC expansion and function, contributing to pancreatic tumor progression. Thus, SHIP-1 can be a potential therapeutic target to help restore immunohomeostasis and improve therapeutic responses in patients with pancreatic cancer.

Hemophagocytic lymphohistiocytosis (HLH) is a hyper-inflammatory clinical syndrome associated with neoplastic disorders especially lymphoma, autoimmune conditions, and infectious agents including bacteria, viruses, protozoa and fungi. In both human and veterinary medicine, hemophagocytic histiocytic disorders are clinically important and frequently fatal. HLH in humans can be a primary (familial, autosomal recessive) or secondary (acquired) condition, with both types generally precipitated by an infectious agent. Previously, no mouse model for secondary HLH has been reported. Using Salmonella enterica serotype Typhimurium by oral gavage to mimic naturally-occurring infection in Sv129S6 mice, we characterized the clinical, hematologic and morphologic host responses to disease thereby describing an animal model with the clinico-pathologic features of secondary HLH as set forth by the Histiocyte Society: fever, splenomegaly, cytopenias (anemia, thrombocytopenia), hemophagocytosis in bone marrow and spleen, hyperferritinemia, and hypofibrinogenemia. Disease severity correlates with high splenic and hepatic bacterial load, and we show disease course can be monitored and tracked in live animals. Whereby secondary HLH is known to occur in human patients with typhoid fever and other infectious diseases, our characterization of a viable natural disease model of secondary HLH offers an important means to elucidate pathogenesis of poorly understood mechanisms of secondary HLH and investigation of novel therapies. We characterize previously unreported secondary HLH in a chronic mouse model of typhoid fever, and novel changes in hematology including decreased tissue ferric iron storage that differs from classically described anemia of chronic disease. Our studies demonstrate S. Typhimurium infection of mice is a natural infectious disease model of secondary HLH that may have utility for elucidating disease pathogenesis and developing novel therapies.

Background: The hypothesis is that hemoglobin-based metrics are useful tools for estimating malaria endemicity and for monitoring malaria control strategies. The aim of this study is to compare population hemoglobin mean and anemia prevalence to established indicators of malaria endemicity, including parasite rates, rates of enlarged spleens in children, and records of (presumptive) malaria diagnosis among populations living with different levels of malaria transmission., Methodology/principal Findings: Convenience sample, multisite cross-sectional household surveys conducted in Papua New Guinea. Correlations (r(2)) between population Hb mean and anemia prevalence and altitude, parasite rate, and spleen rate were investigated in children ages 2 to 10 years, and in the general population; 21,664 individuals from 156 different communities were surveyed. Altitude ranged from 5 to 2120 meters. In young children, correlations between altitude and parasite rate, population Hb mean, anemia prevalence, and spleen rate were high (r(2): -0.77, 0.73, -0.81, and -0.68; p<0.001). In the general population, correlations between altitude and population Hb mean and anemia prevalence were 0.83 and 0.85, respectively. Among young children, parasite rate correlated highly with anemia prevalence, population Hb mean, and spleen rate (r(2): 0.81, -0.81, and 0.86; p<0.001). Population Hb mean (corrected for direct altitude effects) increased with altitude, from 10.5 g/dl at <500 m to 12.8 g/dl at >1500 m (p<0.001)., Conclusions/significance: In PNG, where Plasmodium vivax accounts for an important part of all malaria infections, population hemoglobin mean and anemia prevalence correlate well with altitude, parasite, and spleen rates. Hb measurement is simple and affordable, and may be a useful new tool, alone or in association with other metrics, for estimating malaria endemicity and monitoring effectiveness of malaria control programs. Further prospective studies in areas with different malaria epidemiology and different factors contributing to the burden of anemia are warranted to investigate the usefulness of Hb metrics in monitoring malaria transmission intensity.