Surface TREM2 on circulating M-MDSCs as a novel prognostic factor for adults with treatment-naïve diffuse large B-cell lymphoma.

INTRODUCTION: Circulating monocytic myeloid-derived suppressive cells (M-MDSCs) are implicated as a poor prognostic factor and cause CAR T-cell failure in diffuse large B-cell lymphoma (DLBCL). Triggering receptors expressed on myeloid cells 2 (TREM2) are a transmembrane glycoprotein that polarize macrophages to anti-inflammation phenotype but have never been explored on M-MDSCs. This study aims to elucidate the expression and clinical impact of surface TREM2 on circulating M-MDSCs derived from DLBCL adults. METHODS: This prospective, observational study enrolled 100 adults with newly diagnosed and treatment-naïve DLBCL from May 2019 to October 2021. Human circulating M-MDSCs were obtained from freshly isolated peripheral blood, and each patient's surface-TREM2 level on M-MDSCs was normalized via a healthy control at the same performance of flow-cytometry analysis. Murine MDSCs derived from bone marrow (BM-MDSCs) were adopted to assess the link between Trem2 and cytotoxic T lymphocytes. RESULTS: More circulating M-MDSCs at diagnosis of DLBCL predicted worse progression-free (PFS) and overall survival (OS). Patients with higher IPI scores, bone marrow involvement, or lower absolute counts of CD4+ or CD8+ T cells in PB had significantly higher normalized TREM2 levels on M-MDSCs. Additionally, normalized TREM2 levels on M-MDSCs could be grouped into low (< 2%), medium (2-44%), or high (> 44%) levels, and a high normalized TREM2 level on M-MDSCs was proven as an independent prognostic factor for both PFS and OS via multivariate Cox regression analysis and associated with worst PFS and OS. Interestingly, normalized levels of surface TREM2 on M-MDSCs were negatively associated with absolute counts of PB CD8+ T cells and positively correlated with levels of intracellular arginase 1 (ARG1) within M-MDSCs. Wild-type BM-MDSCs had significantly higher mRNA levels of Arg1 and showed more prominent ability to suppress the proliferation of co-cultured CD8+ T cells than BM-MDSCs from Trem2 knockout mice, and the suppressive ability could be impaired by adding Arg1 inhibitors (CB1158) or supplementing L-arginine. CONCLUSION: In treatment-naïve DLBCL adults, a high surface-TREM2 level on circulating M-MDSCs is a poor prognostic factor for both PFS and OS and warrants further investigation for its potential as a novel target in immunotherapy.

LRP-1 links post-translational modifications to efficient presentation of celiac disease-specific T cell antigens.

Celiac disease (CeD) is an autoimmune disorder in which gluten-derived antigens trigger inflammation. Antigenic peptides must undergo site-specific deamidation to be presentable to CD4+ T cells in an HLA-DQ2 or -DQ8 restricted manner. While the biochemical basis for this post-translational modification is understood, its localization in the patient's intestine remains unknown. Here, we describe a mechanism by which gluten peptides undergo deamidation and concentration in the lysosomes of antigen-presenting cells, explaining how the concentration of gluten peptides necessary to elicit an inflammatory response in CeD patients is achieved. A ternary complex forms between a gluten peptide, transglutaminase-2 (TG2), and ubiquitous plasma protein α2-macroglobulin, and is endocytosed by LRP-1. The covalent TG2-peptide adduct undergoes endolysosomal decoupling, yielding the expected deamidated epitope. Our findings invoke a pathogenic role for dendritic cells and/or macrophages in CeD and implicate TG2 in the lysosomal clearance of unwanted self and foreign extracellular proteins.

MGE-Like Neural Progenitor Cell Survival and Expression of Parvalbumin and Proenkephalin in a Jaundiced Rat Model of Kernicterus.

Kernicterus is a permanent condition caused by brain damage from bilirubin toxicity. Dystonia is one of the most debilitating symptoms of kernicterus and results from damage to the globus pallidus (GP). One potential therapeutic strategy to treat dystonia in kernicterus is to replace lost GP neurons and restore basal ganglia circuits through stem cell transplantation. Toward this end, we differentiated human embryonic stem cells (hESCs) into medial ganglion eminence (MGE; the embryological origin of most of the GP neurons)-like neural precursor cells (NPCs). We determined neurochemical phenotype in cell culture and after transplanting into the GP of jaundiced Gunn rats. We also determined grafted cell survival as well as migration, distribution, and morphology after transplantation. As in the GP, most cultured MGE-like NPCs expressed γ-aminobutyric acid (GABA), with some co-expressing markers for parvalbumin (PV) and others expressing markers for pro-enkephalin (PENK). MGE-like NPCs survived in brains at least 7 weeks after transplantation, with most aggregating near the injection site. Grafted cells expressed GABA and PV or PENK as in the normal GP. Although survival was low and the maturity of grafted cells varied, many cells produced neurite outgrowth. While promising, our results suggest the need to further optimize the differentiation protocol for MGE-like NPC for potential use in treating dystonia in kernicterus.

Intrinsic Aerobic Capacity Affects Hippocampal pAkt and HSP72 Response to an Acute High Fat Diet and Heat Treatment in Rats.

BACKGROUND: Aerobic capacity is associated with metabolic, cardiovascular, and neurological health. Low-capacity runner (LCR) rats display low aerobic capacity, metabolic dysfuction, and spatial memory deficits. A heat treatment (HT) can improve metabolic dysfunction in LCR peripheral organs after high fat diet (HFD). Little is known about metabolic changes in the brains of these rats following HT. OBJECTIVE: Our objective was to examine the extent to which high or low aerobic capacity impacts Akt (a protein marker of metabolism) and heat shock protein 72 (HSP72, a marker of heat shock response) after HFD and HT in hippocampus. METHODS: We measured phosphorylated Akt (pAkt) in the striatum and hippocampus, and HSP72 in the hippocampus, of HFD-fed and chow-fed LCR and high-capacity runner (HCR) rats with and without HT. RESULTS: pAkt was lower in the hippocampus of chow-fed LCR than HCR rats. HFD resulted in greater pAkt in LCR but not HCR rats, but HT resulted in lower pAkt in the LCR HFD group. HSP72 was greater in both HCR and LCR rat hippocampus after HT. The HFD blunted this effect in LCR compared to HCR hippocampus. CONCLUSION: The abnormal phosphorylation of Akt and diminished HSP response in the hippocampus of young adult LCR rats might indicate early vulnerability to metabolic challenges in this key brain region associated with learning and memory.

Roles of vitamin A in the regulation of fatty acid synthesis.

Dietary macronutrients and micronutrients play important roles in human health. On the other hand, the excessive energy derived from food is stored in the form of triacylglycerol. A variety of dietary and hormonal factors affect this process through the regulation of the activities and expression levels of those key player enzymes involved in fatty acid biosynthesis such as acetyl-CoA carboxylase, fatty acid synthase, fatty acid elongases, and desaturases. As a micronutrient, vitamin A is essential for the health of humans. Recently, vitamin A has been shown to play a role in the regulation of glucose and lipid metabolism. This review summarizes recent research progresses about the roles of vitamin A in fatty acid synthesis. It focuses on the effects of vitamin A on the activities and expression levels of mRNA and proteins of key enzymes for fatty acid synthesis in vitro and in vivo. It appears that vitamin A status and its signaling pathway regulate the expression levels of enzymes involved in fatty acid synthesis. Future research directions are also discussed.

An oral absorbent, AST-120, restores vascular growth and blood flow in ischemic muscles in diabetic mice via modulation of macrophage transition.

Background Diabetes has a pronounced effect on the peripheral vasculature. The accumulation of advanced glycation end products (AGEs) is regarded as the crucial mechanism responsible for vascular damage in diabetes, but it is not easy to be avoided from food. In this study, we aimed to investigate the effects of an oral absorbent, AST-120, on the accumulation of AGEs and changes in blood flow recovery in diabetic mice. Methods The mice were divided into four groups, wild-type (WT) mice without treatment, WT mice treated with 5% AST-120 mixed into pulverized chow, streptozotocin-induced diabetes mellitus (DM) mice, and DM mice treated with 5% AST-120. Six weeks after hind-limb ischemia surgery, blood flow reperfusion, histology, plasma AGE, and cytokine were examined. Bone marrow cells were cultured and derived into macrophages to evaluate the effects of AGEs on macrophage polarization. Results Plasma AGEs were significantly increased in diabetic mice. AST-120 could bind to AGEs and reduced their plasma concentrations. Histological analysis revealed fewer collateral vessels with corresponding impairment of blood flow recovery in diabetic mice. In these mice, AGE-positive and AGE receptor-positive macrophages were numerous in ischemic limbs compared with non- diabetic mice. In diabetic mice, macrophages in ischemic tissues demonstrated greater M1 polarization than M2 polarization; this pattern was reversed in the AST-120 treatment group. The change in macrophage polarization was associated with the corresponding expression of pro-inflammatory cytokines in the ischemic tissues. In cell cultures, AGEs triggered the transformation of bone marrow-derived macrophages into the M1 phenotype. The alterations in the polarization of macrophages were reversed after treatment with AST-120. Conclusions Oral administration of AST-120 decreased the serum levels of AGEs in diabetic mice and improved neovascularization of ischemic limbs. This benefit may be due to, at least partially, the alterations in macrophage polarization and the associated changes in inflammatory cytokines.

Upregulation of RelB in the miR-122 knockout mice contributes to increased levels of proinflammatory chemokines/cytokines in the liver and macrophages.

OBJECTIVE: MicroRNA-122 (miR-122) is the most abundant miRNA in the liver and it plays an important role in regulating liver metabolism and tumor formation. Previous studies also reveal an anti-inflammatory function of miR-122; however, relatively little is known about the mechanisms by which miR-122 suppresses inflammation. This study aims to search the effect of miR-122 on proinflammatory chemokines/cytokines production in mice. METHODS: Quantitative real-time PCR, Western blot analysis, and ELISA were performed to examine gene expression. TargetScan, miRanda, and microT v3.0 were used to search for possible miR-122 target sites in the 3'-untranslated regions (3'-UTR) of candidate genes. Luciferase reporter assay and site-directed mutagenesis were applied to verify miR-122 target sequences. LPS was applied to peritoneal macrophages and mice to evaluate inflammatory response. RESULTS: The expression of proinflammatory chemokines, including Ccl2, Ccl4, Ccl20, Cxcl2, and Cxcl10, and Relb in the livers of miR-122 knockout (KO) mice was increased. We identified Relb as a direct miR-122 target. Overexpressing RelB in the mouse liver increased the expression of Ccl2, Ccl4, Ccl20, Cxcl2, and Cxcl10. Peritoneal macrophages from miR-122 KO mice had a higher level of RelB, and they showed a stronger NF-κB activation and more TNF-α and IL-6 secretion after LPS stimulation. Overexpression of RelB in a macrophage cell line augmented LPS-induced TNF-α and IL-6 production. miR-122 KO mice showed a greatly increased mortality rate and generated a stronger and lasting inflammatory response to LPS. CONCLUSIONS: Deletion of miR-122 caused an upregulation of proinflammatory chemokines and RelB in the liver. Increased RelB may contribute to increases in these chemokine in the liver. Intriguingly, deletion of miR-122 also enhanced the sensitivity of macrophages and mice to LPS. Our results reveal that reducing RelB expression is a new mechanism by which miR-122 regulates inflammation.

TREM-1-dependent M1 macrophage polarization restores intestinal epithelium damaged by DSS-induced colitis by activating IL-22-producing innate lymphoid cells.

BACKGROUND: Triggering receptor expressed on myeloid cells-1 (TREM-1) is highly expressed on macrophages in inflamed intestines and reportedly promotes inflammatory bowel disease (IBD) by augmenting pro-inflammatory responses. To study the mechanism mediated by TREM-1 on macrophages, we generated an independent TREM-1 deficient mouse. METHODS: Acute colitis was induced in C57BL/6 and TREM-1-deficient mice by the administration of dextran sodium sulfate (DSS). Colonic lamina propria immune cell composition and cytokines were analyzed. An innate lymphoid cell (ILC) co-culture experiment with macrophages was used to analyze IL-22 levels. Exogenous IL-22 and TREM-1-expressing macrophages were supplied to TREM-1-deficient mice for examining their effects on intestinal barrier integrity. RESULTS: In inflamed colons, TREM-1 loss compromised the activation of ILC3 and their production of IL-22, which is required for intestinal barrier integrity. ILC3-mediated IL-22 production depends on IL-1ß secreted by M1-polarized macrophages, and we found that TREM-1 deficiency results in a decreased number of IL-1ß producing-M1 macrophages in colons exposed to DSS. Accordingly, DSS-mediated damage was ameliorated by supplying exogenous IL-22 and TREM-1-expressing macrophages to TREM-1-deficient mice. CONCLUSIONS: TREM-1 plays a crucial role in regulating IL-22 production by ILC3 through modulating M1-macrophage polarization during DSS-induced acute colitis.

Short Term Development and Fate of MGE-Like Neural Progenitor Cells in Jaundiced and Non-Jaundiced Rat Brain.

Neonatal hyperbilirubinemia targets specific brain regions and can lead to kernicterus. One of the most debilitating symptoms of kernicterus is dystonia, which results from bilirubin toxicity to the globus pallidus (GP). Stem cell transplantation into the GP to replace lost neurons and restore basal ganglia circuits function is a potential therapeutic strategy to treat dystonia in kernicterus. In this study we transplanted human medial ganglionic eminence (MGE)-like neural progenitor cells (NPCs) that we differentiated into a primarily gamma-aminobutyric acid (GABA)ergic phenotype, into the GP of non-immunosuppressed jaundiced (jj) and non-jaundiced (Nj) rats. We assessed the survival and development of graft cells at three time-points post-transplantation. While grafted MGE-like NPCs survived and generated abundant fibers in both jj and Nj brains, NPC survival was greater in the jj brain. These results were consistent with our previous finding that excitatory spinal interneuron-like NPCs exhibited a higher survival rate in the jj brain than in the Nj brain. Our findings further support our hypothesis that slightly elevated bilirubin levels in the jj brain served as an antioxidant and immunosuppressant to protect the transplanted cells. We also identified graft fibers growing toward brain regions that receive projections from the GP, as well as host fibers extending toward the graft. These promising findings suggest that MGE-like NPCs may have the capacity to restore the circuits connecting GP and other nuclei.

Region-specific differences in bioenergetic proteins and protein response to acute high fat diet in brains of low and high capacity runner rats.

Aerobic capacity is a strong predictor of mortality. Low capacity runner (LCR) rats exhibit reduced mitochondrial function in peripheral organs. A high fat diet (HFD) can worsen metabolic phenotype in LCR rats. Little is known about metabolic changes in the brains of these rats, however. This study examined protein markers of mitochondrial function and metabolism as a function of aerobic running capacity and an acute HFD in four brain regions: the striatum, hippocampus, hypothalamus, and substantia nigra. After 3 days HFD or chow diets, we measured peroxisome proliferator-activated receptor-γ coactivator 1α (PGC1-α), nuclear respiratory factors 1 (Nrf-1), mitochondrial transcription factor A (TFAM), and phosphorylated (activated) AMP-activated protein kinase (p-AMPK) protein levels in the four brain regions. LCR rats exhibited lower levels of mitochondrial proteins (PGC1-α, Nrf-1, TFAM), and greater p-AMPK, in striatum, but not in the other brain regions. Mitochondrial protein levels were greater in HFD LCR striatum, while p-AMPK was lower in this group. Markers of lower mitochondrial biogenesis and increased metabolic demand were limited to the LCR striatum, which nevertheless maintained the capacity to respond to an acute HFD challenge.

Fate of Neural Progenitor Cells Transplanted Into Jaundiced and Nonjaundiced Rat Brains.

High levels of bilirubin in infants can cause kernicterus, which includes basal ganglia damage and dystonia. Stem cell transplantation may be an effective treatment for this disease. In this study, we transplanted human neural progenitor cells differentiated toward propriospinal interneurons into the striatum of 20-day-old spontaneously jaundiced (jj) Gunn rats and nonjaundiced (Nj) littermates. Using immunohistochemical methods, we found that grafted cells survived and grew fibers in jj and Nj brains 3 weeks after transplantation. Grafted cells had a higher survival rate in jj than in Nj brains, suggesting that slightly elevated bilirubin may protect graft survival due to its antioxidative and immunosuppressive effects. Despite their survival, only a small portion of grafted neurons expressed GAD-6 or ChAT, which mark GABAergic and cholinergic neurons, respectively, and are the cells that we are attempting to replace in kernicterus. Thus, NPCs containing large populations of GABAergic and cholinergic neurons should be used for further study in this field.

Mining Health Social Media with Sentiment Analysis.

With the rapid development of the Internet, more and more users utilize health communities (known as forums) to find health-related information, share their medical stories and experiences, or interact with other people in the communities. In this paper, we propose a framework to analyze the user-generated contents in a health community. The proposed framework contains three phases. First, we extract medical terms, including conditions, symptoms, treatments, effectiveness and side effects to form a virtual document for each question in the community. Next, we modify Latent Dirichlet Allocation (LDA) by adding a weighted scheme, called conLDA, to cluster virtual documents with similar medical term distributions into a conditional topic (C-topic). Finally, we analyze the clustered C-topics by sentiment polarities, and physiological and psychological sentiment. The experiment results show that conLDA outperforms the original LDA, and can cluster relevant medical terms and relevant questions together. The C-topics clustered by conLDA are more thematic than those clustered by the original LDA. The results of sentiment analysis may provide a quick reference and valuable insights for patients, caregivers and doctors.

C5L2 is required for C5a-triggered receptor internalization and ERK signaling.

C5L2 is a receptor that binds to C5a and belongs to the family of G protein-coupled receptors, but its role in physiological C5a-mediated responses remains under debate. Here we show that, like the canonical C5a receptor C5aR, C5L2 plays a pro-inflammatory role in a murine model of acute experimental colitis. We demonstrate that C5L2 physically interacts with C5aR and is required for optimal C5a-mediated C5aR internalization and associated ERK activation. Abrogation of C5a-induced receptor internalization by treatment with the dynamin inhibitor dynasore(TM) impaired C5a-induced MEK and ERK signaling. Although the presence of C5aR alone was sufficient to recruit the scaffold protein ß-arrestin1 to the cell membrane in response to C5a stimulation, it was inadequate to mediate AP2 recruitment and subsequent C5aR internalization. Expression of C5L2 allowed normal internalization of C5aR in response to C5a stimulation, followed by normal ERK signaling. Thus, our work reveals an essential role for C5L2 in C5a-triggered, AP2-dependent C5aR internalization and downstream ERK signaling.

TREM-1 promotes survival during Klebsiella pneumoniae liver abscess in mice.

Klebsiella pneumoniae liver abscess (KPLA) is prevalent in East Asia. Liver abscess can develop after translocation of K. pneumoniae from a patient's bowel into the liver via the portal circulation. TREM-1 (triggering receptor expressed on myeloid cells 1) amplifies inflammatory signaling during infection, but its role in KPLA is poorly understood. We used an animal study to characterize the role of TREM-1 in KPLA. We compared survival rates, bacterial burdens in tissues, inflammatory cytokine levels, and histology findings between wild-type and Trem-1 knockout (KO) mice after oral inoculation of capsular type K1 K. pneumoniae. Translocation of K. pneumoniae to mesenteric lymph nodes and liver was examined, and intestinal permeability, antimicrobial peptide expression, and the clearance of K. pneumoniae in the small intestine were determined. In the absence of TREM-1, KPLA model mice showed increased K. pneumoniae dissemination, enhanced liver and systemic inflammation, and reduced survival. Impaired bacterial clearance in the small intestine causes enhanced K. pneumoniae translocation, which renders Trem-1 KO mice more susceptible to K. pneumoniae oral infection. In conclusion, TREM-1-mediated bacterial clearance in the small intestine is an important immune response against K. pneumoniae. TREM-1 deficiency enhances K. pneumoniae translocation in the small intestine and increases mortality rates in mice with KPLA.

Fungal polymerase chain reaction testing in equine ulcerative keratitis.

OBJECTIVE: To assess the diagnostic utility of fungal polymerase chain reaction (PCR) in forty-three horses with naturally acquired corneal ulcers presenting to a private practice. METHODS: Routine evaluation of cytologic, histologic, and microbiologic samples was performed. Two PCR approaches were compared - generic and specific fungal nested PCR followed by sequencing and quantitative PCR (qPCR). PCRs were applied to pure control fungal cultures, corneal tissue from ulcerated eyes and in a subset of 9 horses, to swabs from contralateral normal eyes. RESULTS: The expected fungus was identified by nested PCR and qPCR in all control fungal cultures. In all fungal culture-positive affected eyes (10/43), one or more fungi were identified by nested PCR and 4/10 were positive by qPCR. In 6/10 animals, the same fungus was identified by nested PCR and culture. Of these 6, only three were positive by qPCR. Fungal agents were identified by morphology in 8/10 horses. Diagnosis of fungal keratitis was reserved for only those cases in which the same fungus could be identified by PCR, culture, and morphology (5 horses). In 33/43 culture-negative affected eyes and in 6/9 unaffected eyes, one or more fungi were identified by nested PCR in 26 samples and by qPCR in 2 samples. Apart from Aspergillus spp, similar fungi were identified in affected and control eyes. Most eyes harbored mixed bacterial and fungal agents. CONCLUSIONS: Nested PCR results confirmed all cytologically positive cases of fungal keratitis. Nested PCR identified a greater spectrum of agents than either culture or qPCR.

Characterization of ertapenem-resistant Enterobacter cloacae in a Taiwanese university hospital.

The emergence of carbapenem resistance in Enterobacteriaceae has become a great concern. The aim of this study was to characterize ertapenem-resistant Enterobacter cloacae isolates from a Taiwanese university hospital. A total of 355 nonduplicated E. cloacae isolates collected in 2007 were analyzed by antimicrobial susceptibility testing with and without an inhibitor of efflux pumps and AmpC ß-lactamase. The phenotype of extended-spectrum ß-lactamase (ESBL), profile of outer membrane proteins (OMPs), and clonal relatedness were investigated by the double-disk synergy test, urea/SDS-PAGE, and pulsed-field gel electrophoresis (PFGE), respectively. ß-Lactamase genes were examined by PCR and sequencing, and the expression of efflux pump gene acrB was evaluated by reverse transcription-PCR. The contribution of porin deficiency to resistance was investigated by restoring functional porin genes on plasmids. We demonstrated that ertapenem resistance was prevalent (53/355; 14.9%) in E. cloacae. Among the strains, IMP-8, SHV-12, and TEM-1 ß-lactamases were identified in 3 (5.7%), 40 (75.5%), and 46 (86.8%) isolates, respectively. PFGE showed clonal diversity among these isolates. Phenotypes of ESBL, AmpC ß-lactamase overproduction, an active efflux pump, and change in the expression of OMPs were found in 18 (34%), 11 (20.8%), 51 (96.2%), and 23 (43.4%) of ertapenem-resistant strains, respectively. Ertapenem MICs were restored in strains with OmpC and OmpF expression plasmids. This study suggests that ESBL, AmpC ß-lactamase overproduction, and decreased OMP expression combined with an active efflux pump contribute to the ertapenem resistance of E. cloacae. The presence of IMP-8 may also play a partial role in ertapenem resistance in Taiwan.