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Nonalcoholic fatty liver disease (NAFLD) is one of the most common liver diseases in adults. NAFLD progresses from benign liver fat accumulation to liver inflammation and cirrhosis, and ultimately leads to liver failure. Although several rodent models have been established for studying NAFLD, they have limitations that include cost, speed of disease development, key dissimilarities, and poor amenability to pharmacological screens. Here, we present a novel 2-hit zebrafish model to replicate aspects of NAFLD pathogenesis. We fed zebrafish larvae a high-fat diet (HFD) to drive liver fat accumulation (first hit). Next, we exacerbated liver-specific inflammation using a transgenic line (fabp10-CETI-PIC3) that induces the expression of proinflammatory cytokines following induction with doxycycline (second hit). These hits promoted fat accumulation and liver inflammation, as demonstrated by the high expression of inflammatory cytokines, macrophage infiltration, stress induction, and hepatic lipid droplet accumulation. Furthermore, zebrafish in this paradigm showed deranged glucose metabolism. To validate a small-molecule screening approach, we treated HFD-fed fish with pioglitazone, a drug shown to be beneficial for NAFLD in humans, and measured a sharp reduction in liver lipid accumulation. These results demonstrate new utility for zebrafish in modeling early NAFLD pathogenesis and demonstrate their feasibility for in vivo screening of new pharmacological interventions.

Adaptive immunity relies on specialized effector functions elicited by lymphocytes, yet how antigen recognition activates appropriate effector responses through nonspecific signaling intermediates is unclear. Here we examined the role of chromatin priming in specifying the functional outputs of effector T cells and found that most of the cis -regulatory landscape active in effector T cells was poised early in development before the expression of the T cell antigen receptor. We identified two principal mechanisms underpinning this poised landscape: the recruitment of the nucleosome remodeler mammalian SWItch/Sucrose Non-Fermentable (mSWI/SNF) by the transcription factors RUNX1 and PU.1 to establish chromatin accessibility at T effector loci; and a ‘relay’ whereby the transcription factor BCL11B succeeded PU.1 to maintain occupancy of the chromatin remodeling complex mSWI/SNF together with RUNX1, after PU.1 silencing during lineage commitment. These mechanisms define modes by which T cells acquire the potential to elicit specialized effector functions early in their ontogeny and underscore the importance of integrating extrinsic cues to the developmentally specified intrinsic program. Koh et al. show that loci active in differentiated effector T cells are poised in early T precursors before the expression of T cell antigen receptors in a manner dependent on the chromatin remodeling complex mammalian SWItch/Sucrose Non-Fermentable and the PU.1–RUNX1 and BCL11B–RUNX1 complexes.

Calculations of gamma-ray mass attenuation coefficients of various detector materials (crystals) were carried out by means of FLUKA Monte Carlo (MC) method at different gamma-ray energies. NaI, PVT, GSO, GaAs and CdWO4 detector materials were chosen in the calculations. Calculated coefficients were also compared with the National Institute of Standards and Technology (NIST) values. Obtained results through this method were highly in accordance with those of the NIST values. It was concluded from the study that FLUKA MC method can be an alternative way to calculate the gamma-ray mass attenuation coefficients of the detector materials.

A Hamamatsu R1828-01 model fast photomultiplier tube, i.e. having fast time response, was tested whether it can be utilized for the detection of alpha particles. An Eljen AJ-440 model ZnS(Ag) scintillator sheet was mounted on the front window of the PMT. In order to obtain best alpha detection performance, three different main amplifier models that are in our laboratory were used. The alpha energy resolution values were calculated through the amplifiers. Thus, the most appropriate amplifier model in the used ones was determined. It was shown that a fast PMT could be used in alpha particle detection in addition to the timing measurement. With this study, a Hamamatsu R18128-01 type PMT was first tested for the alpha detection and the detection performances through the main amplifiers were compared.

Disclosure: E. Ermis: None. T. Nargis: None. A. Kulkarni: None. S. May: None. S.A. Tersey: None. R.G. Mirmira: None. R.M. Anderson: None. Type 1 diabetes (T1D) is an autoimmune disease, characterized in part by islet inflammation and macrophage infiltration leading to β-cell injury and loss. The inflammatory mediator 12/15-lipoxygenase (12/15-LOX) and its lipid product 12-hydroxyeicosatetraenoic acid (12-HETE) have both been implicated in T1D. 12-HETE activates the low affinity receptor leukotriene B4 receptor 2 (LTB4R2, or simply BLT2), which was reported as a low-affinity receptor for 12-HETE, suggesting that BLT2 may play a proinflammatory role. Here, we used both mouse and zebrafish models to probe the roles of BLT2 in macrophage activation and/or recruitment to pancreatic islets during T1D progression. First, we analyzed Blt2 gene expression in a variety of C57BL/6J mouse tissues. We found Blt2 gene expression in bone marrow derived macrophages and peritoneal macrophages, however we found little to no expression in pancreatic islets. Although Blt2-/- mice showed a significantly reduced expression of Blt2, they exhibited overall normal growth and glucose tolerance. To uncover roles for BLT2 in macrophage function, we first interrogated its involvement in the in vitro polarization of proinflammatory macrophages to the M1-like state using LPS and IFN-γ and the M2-like state using IL-4. Under these conditions, we found no differences between Blt2-/- and wildtype macrophages, as assessed by flow cytometry analysis, suggesting that BLT2 does not play a substantial role in macrophage polarization. Next, to query roles for BLT2 in macrophage migration, we employed the transgenic zebrafish line Tg(mpeg:EGFP) in which macrophages are labeled with green fluorescent protein (GFP). Following zebrafish tailfin injury, we quantified macrophage accumulation at the injury at 6 hours post amputation. Inhibitory treatments of zebrafish larvae with either a chemical BLT2 inhibitor (LY255283) or by knockdown of the blt2a gene using an antisense morpholino, both decreased the number of GFP+ macrophages found at the injury site. Moreover, in the latter approach ectopic expression of blt2a mRNA effectively restored the wild-type phenotype. To determine if BLT2 mediates macrophage migration to injured islets, we employed a transgenic zebrafish model of β-cell injury, Tg(ins:NTR), in which metronidazole treatment rapidly induces β-cell injury. We treated the Tg(mpeg: GFP); Tg(ins: NTR) zebrafish with metronidazole and monitored macrophage migration into injured islets. We observed a significant decrease in macrophage migration towards injured β-cells in blt2a-knockdown zebrafish. Collectively, these results show that BLT2 plays a role in macrophage migration associated with tissue injury, including the islet β cell as seen in T1D. Presentation: Saturday, June 17, 2023

The patella plays a vital role in stifle joint function by contributing to limb extension and joint stabilization. While its clinical significance in small animal orthopedic surgery is well established, detailed comparative data on patellar morphology in domestic cats and dogs remain limited. This study aimed to investigate interspecific and intraspecific variation in patellar shape and size using three-dimensional geometric morphometric techniques. Computed tomography images of 18 cats and 55 dogs were used to construct 3D models, and a total of 14 anatomical landmarks were manually placed on each patella. Generalized Procrustes Analysis was applied, followed by principal component analysis to explore patterns of shape variation. The results revealed a significant difference in shape between cats and dogs, with dogs exhibiting broader variation and larger centroid sizes. Regression analysis indicated that 12.2% of the observed shape variation could be attributed to centroid size, reflecting the influence of patellar size. This strong link suggests that size has a significant influence on how the patella’s shape varies, especially in dogs. These findings highlight the importance of individual variation in patellar morphology, particularly in dogs. A single standardized implant or surgical technique may not be appropriate for all cases. Integrating shape and size evaluations into preoperative imaging can help improve implant fit, joint stability, and overall surgical success. In the absence of breed-specific data, individualized planning remains the most reliable approach.

Background: Caspase-3 is involved in the execution of apoptosis and is widely used as an apoptotic marker. Tumor necrosis factor-α(TNF-α) released from activated macrophages has various functions such as modulation of cell growth and differentiation, immunoregulation, coagulation, and regulation of endothelial cell function. This study investigated the immunohistochemical staining of caspase-3 and TNF-αexpression in the placentas of pregnant women with preterm premature rupture of membranes (PPROM). Methods: Placentas of 25 healthy, and 25 women with PPROM were processed for routine histological tissue processing. Placentas were stained with hematoxylin-eosin, caspase-3, and TNF-αimmunostaining. Results: Normal placental histology was observed in the control group. Amniotic epithelium, vascular structures, and fibrinoid accumulation were histologically normal. Leukocyte infiltration, thinned vessel walls with dilatation and congestion, syncytial nodes, and fibrinoid accumulation were increased in the PPROM group. The immune activity of caspase-3 expression was mainly negative in placental components such as syncytial nodes, vascular endothelium, fibrinoid accumulation, and macrophages in the control group. In the PPROM group, caspase-3 positive reaction was increased in the amniotic membrane and epithelium, endothelial cells, fibrinoid accumulation, and areas of inflammatory cell infiltration. In the control group, negative TNF-αexpression was observed in the placental membranes and structures. In the PPROM group, TNF-αexpression was increased in inflammatory cells, endothelial cells, and syncytial nodes. Conclusions: Placentas of patients with PPROM showed loss and weakened membranes with increased placental pathology, and increased expression of caspase-3 and TNF-α. We suggest that caspase-3 and TNF-αsignaling pathways can be used as a marker in the progression of PPROM.

The present study aimed to identify the frequency of Fabry disease in patients with cardiac hypertrophy of unknown etiology and to evaluate demographic and clinical characteristics, enzyme activity levels, and genetic mutations at the time of diagnosis. This national, multicenter, cross-sectional, single-arm, observational registry study was conducted in adult patients with a clinical echocardiographic diagnosis of left ventricular hypertrophy and/or the presence of prominent papillary muscle. In both genders, genetic analysis was performed by DNA Sanger sequence analysis. A total of 406 patients with left ventricular hypertrophy of unknown origin were included. Of the patients, 19.5% had decreased enzyme activity (≤2.5 nmol/mL/h). Although genetic analysis revealed GLA (galactosidase alpha) gene mutation in only 2  patients (0.5%), these patients were considered to have probable but not 'definite Fabry disease' due to normal lyso Gb3 levels and gene mutations categorized as variants of unknown significance. The prevalence of Fabry disease varies according to the characteristics of the population screened and the definition of the disease used in these trials. From cardiology perspective, left ventricular hypertrophy is the major reason to consider screening for Fabry disease. Enzyme testing, genetic analysis, substrate analysis, histopathological examination, and family screening should be performed, when necessary, for a definite diagnosis of Fabry disease. The results of this study underline the importance of the comprehensive use of these diagnostic tools to reach a definite diagnosis. The diagnosis and management of Fabry disease should not be based solely on the results of the screening tests.

The aim of the present study was to evaluate the efficacy and safety of eltrombopag, an oral thrombopoietin receptor agonist, in patients with chronic immune thrombocytopenia (ITP). A total of 285 chronic ITP patients (187 women, 65.6%; 98 men, 34.4%) followed in 55 centers were enrolled in this retrospective cohort. Response to treatment was assessed according to platelet count (/mm ) and defined as complete (platelet count of >100,000/mm ), partial (30,000-100,000/mm or doubling of platelet count after treatment), or unresponsive (<30,000/mm ). Clinical findings, descriptive features, response to treatment, and side effects were recorded. Correlations between descriptive, clinical, and hematological parameters were analyzed. The median age at diagnosis was 43.9±20.6 (range: 3-95) years and the duration of follow-up was 18.0±6.4 (range: 6-28.2) months. Overall response rate was 86.7% (n=247). Complete and partial responses were observed in 182 (63.8%) and 65 (22.8%) patients, respectively. Thirty-eight patients (13.4%) did not respond to eltrombopag treatment. For patients above 60 years old (n=68), overall response rate was 89.7% (n=61), and for those above 80 years old (n=12), overall response rate was 83% (n=10). Considering thrombocyte count before treatment, eltrombopag significantly increased platelet count at the 1 , 2 , 3 , 4 , and 8 weeks of treatment. As the time required for partial or complete response increased, response to treatment was significantly reduced. The time to reach the maximum platelet levels after treatment was quite variable (1-202 weeks). Notably, the higher the maximum platelet count after eltrombopag treatment, the more likely that side effects would occur. The most common side effects were headache (21.6%), weakness (13.7%), hepatotoxicity (11.8%), and thrombosis (5.9%). Results of the current study imply that eltrombopag is an effective therapeutic option even in elderly patients with chronic ITP. However, patients must be closely monitored for response and side effects during treatment. Since both response and side effects may be variable throughout the follow-up period, patients should be evaluated dynamically, especially in terms of thrombotic risk factors.