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Dysfunctional phagocytic clearance of β-amyloid (Aβ) in microglia and peripheral macrophages/monocytes has been implicated in Alzheimer's disease (AD), but the mechanisms underlying this dysfunction are not yet well understood. In this study, we examined the role of glia maturation factor-γ (GMFG), an actin-disassembly protein that is highly expressed in immune cells, in macrophage Aβ phagocytosis and in regulating scavenger receptor AI (SR-AI), a cell-surface receptor that has previously been implicated in Aβ clearance. GMFG knockdown increased phagocytosis of Aβ42 in BMDMs and RAW264.7 murine macrophages, while GMFG overexpression reduced Aβ42 uptake in these cells. Blocking with anti-SR-AI antibodies inhibited Aβ42 uptake in GMFG-knockdown cells, establishing a role for SR-AI in Aβ42 phagocytosis. GMFG knockdown increased SR-AI protein expression under both basal conditions and in response to Aβ42 treatment via both the transcriptional and post-transcriptional level in RAW264.7 macrophages. GMFG knockdown modulated Aβ42-induced K48-linked and K63-polyubiquitination of SR-AI, the phosphorylation of SR-AI and JNK, suggesting that GMFG plays a role for intracellular signaling in the SR-AI-mediated uptake of Aβ. Further, GMFG-knockdown cells displayed increased levels of the transcriptional factor MafB, and silencing of MafB in these cells reduced their SR-AI expression. Finally, GMFG was found to interact with the nuclear pore complex component RanBP2, and silencing of RanBP2 in GMFG-knockdown cells reduced their SR-AI expression. Collectively, these data support the role of GMFG as a novel regulator of SR-AI in macrophage Aβ phagocytosis, and may provide insight into therapeutic approaches to potentially slow or prevent the progression of AD., (Published by Oxford University Press on behalf of Society for Leukocyte Biology 2024.)

There are ∼4500 genes within the 'druggable genome', the subset of the human genome that expresses proteins able to bind drug-like molecules, yet existing drugs only target a few hundred. A substantial subset of druggable proteins are largely uncharacterized or understudied, with many falling within G protein-coupled receptor (GPCR), ion channel, and kinase protein families. To improve scientific understanding of these three understudied protein families, the US National Institutes of Health launched the Illuminating the Druggable Genome Program. Now, as the program draws to a close, this review will lay out resources developed by the program that are intended to equip the scientific community with the tools necessary to explore previously understudied biology with the potential to rapidly impact human health., (Copyright © 2023. Published by Elsevier Ltd.)

Olfactomedin4 (Olfm4) is expressed in normal mouse prostate. However, Olfm4+ cells in the murine prostate have not been well characterized. In this study, we generated an Olfm4 eGFP reporter mouse line with C57BL/6 mice and investigated the distribution of Olfm4/eGFP-expressing cells during postnatal development from P1, P7, P14, P20, P42, P56 to adult male mouse prostate and urethral tube. We observed Olfm4/eGFP expression in urogenital and prostatic epithelial cells during early postnatal development, which persisted into adulthood in urethral-tube and anterior-prostate (AP) epithelium. We found Olfm4+ cells are E-cadherin+/CD44+/Foxa1+ and some of subpopulation are Ck8+/Ck5+/Sca-1-/Ck4-/Syn- in the adult mouse AP epithelium. Functional studies of single-cell preparations of Olfm4/eGFP-expressing cells isolated from adult Olfm4 eGFP mouse prostate demonstrated that Olfm4+ cells can grow and form colonies, spheres, or organoids in culture. Bioinformatic analysis of Olfm4+ cells using single-cell RNA sequencing meta data in adult mouse urethra (GSE145865) identified upregulation of genes related to cell and tissue migration and development, as well as upregulation of xenobiotic metabolism signaling pathways. In conclusion, Olfm4 eGFP mouse is a novel model to further study Olfm4's biological functions and Olfm4+ cells may contribute importantly to cellular processes supporting development and homeostasis of the epithelium in murine prostate and urethral tube., (© 2023. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Sickle cell disease (SCD) and β-thalassemia are among the most common genetic disorders worldwide, affecting global health and mortality. Hemoglobin A2 (HbA2, α2δ2) is expressed at a low level in adult blood due to the lack of the Kruppel-like factor 1 (KLF1) binding motif in the δ-globin promoter region. However, HbA2 is fully functional as an oxygen transporter, and could be a valid antisickling agent in SCD, as well as a substitute for hemoglobin A in β-thalassemia. We have previously demonstrated that KLF1-GATA1 fusion protein could interact with the δ-globin promoter and increase δ-globin expression in human primary CD34+ cells. We report the effects of 2 KLF1-GATA1 fusion proteins on hemoglobin expression, as well as SCD phenotypic correction in vitro and in vivo. Forced expression of KLF1-GATA1 fusion protein enhanced δ-globin gene and HbA2 expression, as well as reduced hypoxia-related sickling, in erythroid cells cultured from both human sickle CD34+ cells and SCD mouse hematopoietic stem cells (HSCs). The fusion proteins had no impact on erythroid cell differentiation, proliferation, and enucleation. Transplantation of highly purified SCD mouse HSCs expressing KLF1-GATA1 fusion protein into SCD mice lessened the severity of the anemia, reduced the sickling of red blood cells, improved SCD-related pathological alterations in spleen, kidney, and liver, and restored urine-concentrating ability in recipient mice. Taken together, these results indicate that the use of KLF1-GATA1 fusion constructs may represent a new gene therapy approach for hemoglobinopathies., (© 2022 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Competing Interests: The authors declare they have no conflicts of interest.

Biomarkers of infectious diseases are essential tools for patient monitoring, diagnostics, and prognostics. Here we review recent advances in our understanding of olfactomedin 4 (OLFM4) in neutrophil biology and of OLFM4 as a new biomarker for certain infectious diseases. OLFM4 is a neutrophil-specific granule protein that is expressed in a subset of human and mouse neutrophils. OLFM4 expression is upregulated in many viral and bacterial infections, as well as in malaria. OLFM4 appears to play an important role in regulating host innate immunity against bacterial infection. Further, higher expression of OLFM4 is associated with severity of disease for dengue virus, respiratory syncytial virus, and malaria infections. In addition, higher expression of OLFM4 or a higher percentage of OLFM4 + neutrophils is associated with poorer outcomes in septic patients. OLFM4 is a promising biomarker and potential therapeutic target in certain infectious diseases., (Published by Oxford University Press on behalf of Infectious Diseases Society of America 2022.)

Small intestine adenocarcinoma is a rare intestinal malignancy with distinct clinical, pathological, and molecular characteristics. Recently, a fusion of the intestinal stem-cell marker olfactomedin 4 (OLFM4) and the proto-oncogene RET has been identified in a small intestine adenocarcinoma patient. Here we investigated the biological effects of OLFM4-RET fusion and whether it can initiate tumorigenesis in small intestine. OLFM4 expression was found to be frequently lost or reduced in human small intestine adenocarcinoma, and its downregulation correlated with high tumor grade and advanced tumor stage. Expression of OLFM4-RET fusion-induced cellular transformation in HEK293 cells and blocked RET-induced inhibition of colony growth in HuTu 80 small intestine adenocarcinoma cells. Further, expression of OLFM4-RET activated the RAS-RAF-MAPK and STAT3 cell signaling pathways in both HEK293 cells and HuTu 80 cells. OLFM4-RET expression in HEK293 cells upregulated multiple families of genes related to carcinogenesis, cancer progression, and metastasis. Targeted expression of OLFM4-RET in the small intestine led to the development of hyperplasia, adenoma, or adenocarcinoma in transgenic mice. Our study suggests that OLFM4-RET is an oncogenic driver of small intestine tumorigenesis. Therefore, the small intestine adenocarcinoma patients with OLFM4-RET fusion may benefit from treatment with RET kinase inhibitor., (© 2021. This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply.)

Chronic metabolic disorders such as diabetes and obesity are major public health issues in the United States. However, significant disparities in their prevalence and incidence place a greater burden on US racial and ethnic minority groups, contributing to worse COVID-19 outcomes in many. Improving treatment and prevention of diabetes and obesity is critical to the NIDDK. In this Perspective, we will review the burden of metabolic diseases in the United States, the observed disparities for metabolic diseases in relation to COVID-19, and research opportunities to address underlying causes of metabolic diseases, their associated health disparities, and COVID-19., Competing Interests: Declaration of interests The authors declare no competing interests., (Published by Elsevier Inc.)

Olfactomedin 4 (OLFM4) is expressed in normal prostate epithelial cells and immortalized normal human prostate epithelial cells (RWPE1), but the identity of OLFM4-expressing cells within these populations and OLFM4's physiological functions in these cells have not been elucidated. Using single-cell RNA sequencing analysis, we found here that OLFM4 was expressed in multiple stem/progenitor-like cell populations in both the normal prostate epithelium and RWPE1 cells and was frequently co-expressed with KRT13 and LY6D in RWPE1 cells. Functionally, OLFM4-knockout RWPE1 cells exhibited enhanced proliferation of the stem/progenitor-like cell population, shifts stem/progenitor-like cell division to favor symmetric division and differentiated into higher levels PSA expression cells in organoid assays compared with OLFM4-wild RWPE1 cells. Bulk-cell RNA sequencing analysis pinpointed that cMYC expression were enhanced in the OLFM4-knockout RWPE1 cells compared with OLFM4-wild cells. Molecular and signaling pathway studies revealed an increase in the WNT/APC/MYC signaling pathway gene signature, as well as that of MYC target genes that regulate multiple biological processes, in OLFM4-knockout RWPE1 cells. These findings indicated that OLFM4 is co-expressed with multiple stem/progenitor cell marker genes in prostate epithelial cells and acts as a novel mediator in prostate stem/progenitor cell proliferation and differentiation.

The olfactomedin 4 (OLFM4) gene has been analyzed as a tumor-suppressor gene and a putative biomarker in many cancers. In our study, we analyzed the relationship of OLFM4 expression with clinicopathological features and with CpG site methylation in the OLFM4 gene promoter region in human primary prostate adenocarcinoma. OLFM4 protein expression was significantly reduced in prostate cancer tissue compared to adjacent normal tissue and was further significantly reduced in more advanced cancers. Bioinformatic studies with clinical datasets revealed that primary prostate adenocarcinoma patients with reduced OLFM4 mRNA expression exhibited higher Gleason scores and higher preoperative serum prostate-specific antigen levels, as well as lower recurrence-free survival. Three of the eight CpG sites in the OLFM4 gene promoter region were hypermethylated in cancerous prostate cells compared to adjacent normal cells, and reduced methylation of eight CpG sites was associated with increased OLFM4 mRNA expression in RWPE1 and PC-3 cells. Furthermore, knockdown of OLFM4 gene expression was associated with enhanced epithelial-mesenchymal transition (EMT)-marker expression in RWPE immortalized normal prostate cells. In contrast, restoration of OLFM4 expression in PC-3 and DU145 prostate cancer cells lacking OLFM4 significantly inhibited both EMT-marker expression and tumor cell growth in in vitro and in vivo models, indicating that OLFM4 may play a tumor-suppressor role in inhibiting the EMT program, as well as tumor initiation and growth, in prostate cells. Taken together, these findings suggest that OLFM4 plays an important tumor-suppressor role in prostate cancer progression and might be useful as a novel candidate biomarker for prostate cancer., (© 2019 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

Background: Benign ethnic neutropenia (BEN), defined by neutrophil count <1.5 k/μL in the absence of other causes, is an asymptomatic condition more commonly observed in individuals of African ancestry. However, the natural history of this condition has been less well described., Methods: Individuals with BEN were retrospectively identified by chart review or referral to hematology clinics. They were then invited to enroll in a prospective natural history study. Retrospective and prospective clinical and laboratory data were combined for descriptive analyses., Findings: 46 participants, younger and older adults from 2 institutions, had BEN. Hypertension was reported in 30%, musculoskeletal disorders in 15%, and upper respiratory infection in 33% of these adults. Their leukopenia resulted from isolated neutropenia, ranging from 1000 and 1500 cells/μL. The severity of infections was mild and the frequency was similar to other healthy individuals in the ambulatory clinic., Interpretation: In this group of BEN participants, their leukopenia was stable over time, and they had low rates of infections or common medical disorders, confirming the benign nature of this condition. The presence of BEN in children, younger adults, and older adults suggest a hereditary pattern for BEN., (Published by Elsevier Inc.)

In macrophages, cellular iron metabolism status is tightly integrated with macrophage phenotype and associated with mitochondrial function. However, how molecular events regulate mitochondrial activity to integrate regulation of iron metabolism and macrophage phenotype remains unclear. Here, we explored the important role of the actin-regulatory protein glia maturation factor-γ (GMFG) in the regulation of cellular iron metabolism and macrophage phenotype. We found that GMFG was downregulated in murine macrophages by exposure to iron and hydrogen peroxide. GMFG knockdown altered the expression of iron metabolism proteins and increased iron levels in murine macrophages and concomitantly promoted their polarization toward an anti-inflammatory M2 phenotype. GMFG-knockdown macrophages exhibited moderately increased levels of mitochondrial reactive oxygen species (mtROS), which were accompanied by decreased expression of some mitochondrial respiration chain components, including the iron-sulfur cluster assembly scaffold protein ISCU as well as the antioxidant enzymes SOD1 and SOD2. Importantly, treatment of GMFG-knockdown macrophages with the antioxidant N-acetylcysteine reversed the altered expression of iron metabolism proteins and significantly inhibited the enhanced gene expression of M2 macrophage markers, suggesting that mtROS is mechanistically linked to cellular iron metabolism and macrophage phenotype. Finally, GMFG interacted with the mitochondrial membrane ATPase ATAD3A, suggesting that GMFG knockdown-induced mtROS production might be attributed to alteration of mitochondrial function in macrophages. Our findings suggest that GMFG is an important regulator in cellular iron metabolism and macrophage phenotype and could be a novel therapeutic target for modulating macrophage function in immune and metabolic disorders.

Neutrophils increase production of reactive oxygen species, including superoxide, hydrogen peroxide (H 2 O 2 ), and hydroxyl radical, to destroy invading microorganisms under pathological conditions. Conversely, oxidative stress conditions, such as the presence of H 2 O 2 , induce neutrophil apoptosis, which helps to remove neutrophils after inflammation. However, the detailed molecular mechanisms that are involved in the latter process have not been elucidated. In this study, we investigated the potential role of olfactomedin 4 (Olfm4) in H 2 O 2 -induced superoxide production and apoptosis in mouse neutrophils. We have demonstrated that Olfm4 is not required for maximal-dosage PMA- and Escherichia coli bacteria-induced superoxide production, but Olfm4 contributes to suboptimal-dosage PMA- and H 2 O 2 -induced superoxide production. Using an NADPH oxidase inhibitor and gp91phox-deficient mouse neutrophils, we found that NAPDH oxidase was required for PMA-stimulated superoxide production and that Olfm4 mediated H 2 O 2 -induced superoxide production through NADPH oxidase, in mouse neutrophils. We have shown that neutrophils from Olfm4-deficient mice exhibited reduced H 2 O 2 -induced apoptosis compared with neutrophils from wild-type mice. We also demonstrated that neutrophils from Olfm4-deficient mice exhibited reduced H 2 O 2 -stimulated mitochondrial damage and membrane permeability, and as well as reduced caspase-3 and caspase-9 activity, compared with neutrophils from wild-type mice. Moreover, the cytoplasmic translocation of the proapoptotic mitochondrial proteins Omi/HtrA2 and Smac/DIABLO in response to H 2 O 2 was reduced in neutrophils from Olfm4-deficient mice compared with neutrophils from wild-type mice. Our study demonstrates that Olfm4 contributes to H 2 O 2 -induced NADPH oxidase activation and apoptosis in mouse neutrophils. Olfactomedin 4 might prove to be a potential target for future studies on inflammatory neutrophil biology and for inflammatory disease treatment.

Glucose-stimulated insulin secretion (GSIS) is essential for blood glucose homeostasis and is impaired in type 2 diabetes mellitus. Understanding the regulatory components of GSIS has clinical implications for diabetes treatment. In this study, we found that olfactomedin 4 (OLFM4) is endogenously expressed in pancreatic islet β cells and further investigated its potential roles in glucose homeostasis and the pathogenesis of type 2 diabetes using mouse models. Olfm4-deficient mice showed significantly improved glucose tolerance and significantly increased insulin levels after glucose challenge compared with wild-type (WT) mice. GSIS, mitochondrial ATP production, and mitochondrial respiration were all significantly increased in islets isolated from Olfm4-deficient mice compared with those isolated from WT mice. In a high-fat diet (HFD)-induced diabetic mouse model, the increase in insulin levels after glucose challenge was significantly higher in Olfm4-deficient mice compared with WT mice. The impaired glucose tolerance and insulin resistance in HFD-fed mice were improved by loss of Olfm4. Olfm4 was found to be mainly localized in the mitochondria and interacts with GRIM-19 (a gene associated with retinoid-interferon mortality) in Min6 pancreatic β cells. Collectively, these studies suggest that Olfm4 negatively regulates GSIS. OLFM4 may represent a potential therapeutic target for impaired glucose tolerance and patients with type 2 diabetes.

N-oxide derivatives 5(a-b), 8(a-b), and 11(a-c) were designed, synthesized and evaluated in vitro and in vivo as potential drugs that are able to ameliorate sickle cell disease (SCD) symptoms. All of the compounds demonstrated the capacity to releasing nitric oxide at different levels ranging from 0.8 to 30.1%, in vivo analgesic activity and ability to reduce TNF-α levels in the supernatants of monocyte cultures. The most active compound (8b) protected 50.1% against acetic acid-induced abdominal constrictions, while dipyrone, which was used as a control only protected 35%. Compounds 8a and 8b inhibited ADP-induced platelet aggregation by 84% and 76.1%, respectively. Both compounds increased γ-globin in K562 cells at 100 μM. The mechanisms involved in the γ-globin increase are related to the acetylation of histones H3 and H4 that is induced by these compounds. In vitro, the most promising compound (8b) was not cytotoxic, mutagenic and genotoxic., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Background: Benign ethnic neutropenia (BEN) is a hematologic condition associated with people of African ancestry and specific Middle Eastern ethnic groups. Prior genetic association studies in large population showed that rs2814778 in Duffy Antigen Receptor for Chemokines (DARC) gene, specifically DARC null red cell phenotype, was associated with BEN. However, the mechanism of this red cell phenotype leading to low white cell count remained elusive., Methods: We conducted an extreme phenotype design genome-wide association study (GWAS), analyzed ~16 million single nucleotide polymorphisms (SNP) in 1,178 African-Americans individuals from the Reasons for Geographic and Racial Differences in Stroke (REGARDS) study and replicated from 819 African-American participants in the Atherosclerosis Risk in Communities (ARIC) study. Conditional analyses on rs2814778 were performed to identify additional association signals on chromosome 1q22. In a separate cohort of healthy individuals with and without BEN, whole genome gene expression from peripheral blood neutrophils were analyzed for DARC., Results: We confirmed that rs2814778 in DARC was associated with BEN (p = 4.09×10-53). Conditioning on rs2814778 abolished other significant chromosome 1 associations. Inflammatory cytokines (IL-2, 6, and 10) in participants in the Howard University Family Study (HUFS) and Multi-Ethnic Study in Atherosclerosis (MESA) showed similar levels in individuals homozygous for the rs2814778 allele compared to others, indicating cytokine sink hypothesis played a minor role in leukocyte homeostasis. Gene expression in neutrophils of individuals with and without BEN was also similar except for low DARC expression in BEN, suggesting normal function. BEN neutrophils had slightly activated profiles in leukocyte migration and hematopoietic stem cell mobilization pathways (expression fold change <2)., Conclusions: These results in humans support the notion of DARC null erythroid progenitors preferentially differentiating to myeloid cells, leading to activated DARC null neutrophils egressing from circulation to the spleen, and causing relative neutropenia. Collectively, these human data sufficiently explained the mechanism DARC null red cell phenotype causing BEN and further provided a biologic basis that BEN is clinically benign.