Electroacupuncture improves repeated social defeat stress-elicited social avoidance and anxiety-like behaviors by reducing Lipocalin-2 in the hippocampus.

BACKGROUND: Post-traumatic stress disorder (PTSD) is a trauma-related disorder that is associated with pro-inflammatory activation and neurobiological impairments in the brain and leads to a series of affective-like behaviors. Electroacupuncture (EA) has been proposed as a clinically useful therapy for several brain diseases. However, the potential role of EA treatment in PTSD and its molecular and cellular mechanisms has rarely been investigated. METHODS: We used an established preclinical social defeat stress mouse model to study whether EA treatment modulates PTSD-like symptoms and understand its underlying mechanisms. To this end, male C57BL/6 mice were subjected to repeated social defeat stress (RSDS) for 6 consecutive days to induce symptoms of PTSD and treated with EA at Baihui (GV 20) and Dazhui (GV 14) acupoints. RESULTS: The stimulation of EA, but not needle insertion at Baihui (GV 20) and Dazhui (GV 14) acupoints effectively improved PTSD-like behaviors such as, social avoidance and anxiety-like behaviors. However, EA stimulation at the bilateral Tianzong (SI11) acupoints did not affect the PTSD-like behaviors obtained by RSDS. EA stimulation also markedly inhibited astrocyte activation in both the dorsal and ventral hippocampi of RSDS-treated mice. Using next-generation sequencing analysis, our results showed that EA stimulation attenuated RSDS-enhanced lipocalin 2 expression in the hippocampus. Importantly, using double-staining immunofluorescence, we observed that the increased lipocalin 2 expression in astrocytes by RSDS was also reduced by EA stimulation. In addition, intracerebroventricular injection of mouse recombinant lipocalin 2 protein in the lateral ventricles provoked social avoidance, anxiety-like behaviors, and the activation of astrocytes in the hippocampus. Interestingly, the overexpression of lipocalin 2 in the brain also altered the expression of stress-related genes, including monoamine oxidase A, monoamine oxidase B, mineralocorticoid receptor, and glucocorticoid receptor in the hippocampus. CONCLUSIONS: This study suggests that the treatment of EA at Baihui (GV 20) and Dazhui (GV 14) acupoints improves RSDS-induced social avoidance, anxiety-like behaviors, astrocyte activation, and lipocalin 2 expression. Furthermore, our findings also indicate that lipocalin 2 expression in the brain may be an important biomarker for the development of PTSD-related symptoms.

Role of lipocalin-2 in extracellular peroxiredoxin 2-induced brain swelling, inflammation and neuronal death.

Peroxiredoxin-2 (PRX-2) is known to be released from erythrocytes and induce brain damage after intracerebral hemorrhage (ICH); lipocalin-2 (LCN-2) is involved in neuroinflammation following ICH. This study examined the role of LCN-2 in PRX-2 induced brain injury and involved three parts. In the first part, adult male C57BL/6 wild-type (WT), LCN-2 heterozygous (LCN-2 HET), and LCN-2 knockout (LCN-2 KO) mice received either an intracaudate injection of recombinant PRX-2 or saline. In the second part, adult male C57BL/6 WT and male LCN-2 KO mice received recombinant PRX-2 with either recombinant mouse LCN-2 protein or control. In the third part, adult female C57BL/6 WT, LCN-2 HET, and LCN-2 KO mice received recombinant PRX-2. Behavioral tests, and T2- and T2*- weighted magnetic resonance imaging was obtained for all mice. Mice were then euthanized, and their brains used for Western blotting, histology and immunohistochemistry. Intracerebral PRX-2 injections resulted in increased expression of LCN-2 protein. PRX-2-induced brain swelling, neutrophil infiltration, microglia/macrophage activation, neuronal cell death, and neurological deficits were reduced in male LCN-2 HET and LCN-2 KO mice (P < 0.01) compared to WT and were exacerbated by exogenous LCN-2 co-injection. Additionally, intracerebral PRX-2 injections caused brain injury and neurological deficits in female WT mice; effects reduced in female LCN-2 KO mice. In conclusion, intracerebral injection of PRX-2 upregulates LCN-2, and LCN-2 is crucial in the effects of PRX-2 on neutrophil infiltration and microglia/macrophage activation, and ultimately brain damage.

The Effects of Human BDH2 on the Cell Cycle, Differentiation, and Apoptosis and Associations with Leukemia Transformation in Myelodysplastic Syndrome.

Iron overload is related to leukemia transformation in myelodysplastic syndrome (MDS) patients. Siderophores help to transport iron. Type 2-hydroxybutyrate dehydrogenase (BDH2) is a rate-limiting factor in the biogenesis of siderophores. Using qRT-PCR, we analyze BDH2mRNA expression in the bone marrow (BM) of 187 MDS patients, 119 de novo acute myeloid leukemia (AML) patients, and 43 lymphoma patients with normal BM. Elevated BDH2mRNA expression in BM is observed in MDS patients (n = 187 vs. 43, normal BM; P = 0.009), and this is related to ferritin levels. Patients with higher BDH2 expression show a greater risk of leukemia progression (15.25% vs. 3.77%, lower expression; P = 0.017) and shorter leukemia-free-survival (medium LFS, 9 years vs. 7 years; P = 0.024), as do patients with a ferritin level ≥350 ng/mL. Additionally, we investigate the mechanisms related to the prognostic ability of BDH2 by using BDH2-KD THP1. The cell cycle analysis, surface markers, and special stain studies indicate that BDH2-KD induces differentiation and decreases the growth rate of THP1 cells, which is associated with the retardation of the cell cycle. Moreover, many genes, including genes related to mitochondrial catabolism, oncogenes, tumor suppressor genes, and genes related to cell differentiation and proliferation influence BDH2-KD THP1 cells. Herein, we demonstrate that BDH2 is involved in cell cycle arrest and the inhibition of differentiation in malignant cells. Furthermore, the high BDH2 expression in MDS patients could be suggestive of a poor prognostic factor. This study provides a foundation for further research on the roles of BDH2 and iron metabolism in the pathogenesis of MDS.

Potential mechanisms and serum biomarkers involved in sex differences in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a disease associated with high mortality, but notable sex differences have been observed between males and females. For this reason, further research on the mechanisms underlying sex differences in PAH is required to better understand and treat the disease. This study mainly focused on gene expression levels to investigate potential differences in the pathogenesis and progression of PAH between the male and female sexes.Sex-specific differentially expressed genes (DEGs) were analyzed using the Gene Expression Omnibus datasets GSE117261 and GSE38267. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were conducted, and a protein-protein interaction (PPI) network was established based on the identified DEGs to predict potential mechanisms involved in the observed sex differences in the pathogenesis of PAH.We identified 26 female- and 53 male-specific DEGs from lung tissue and 498 female-specific DEGs in blood samples. No male-specific DEGs were identified from blood samples. GO and KEGG pathway analyses revealed that female-specific DEGs in lung tissue were enriched in inflammatory response and cytokine-cytokine receptor interaction, whereas male-specific DEGs were mainly enriched in cellular chemotaxis and the nuclear factor of kappa light polypeptide gene enhancer in B-cell (NF-kappa B) signaling pathway. Lipocalin 2 (LCN2) was the only gene that was differentially expressed in both the lung tissue and the blood of female patients.In conclusion, inflammation and immunity may play key roles in the pathogenesis of female PAH, and LCN2 may act as a serum biomarker of female PAH, whereas the pathogenesis in males is more complicated.

Iron chelators inhibit amyloid-ß-induced production of lipocalin 2 in cultured astrocytes.

Lipocalin 2 (Lcn2) has been implicated to play a role in various neurodegenerative diseases, and normalizing its overexpression may be of therapeutic potential. Iron chelators were found to reduce Lcn2 levels in certain animal models of CNS injury. Focusing on Alzheimer's disease (AD), we found that the iron chelators deferoxamine and deferiprone inhibited amyloid-ß (Aß)-induced Lcn2 production in cultured primary astrocytes. Accordingly, Aß-exposure increased astrocytic ferritin production, indicating the possibility that Aß induces iron accumulation in astrocytes. This effect was not significantly modulated by Lcn2. Known neuroprotective effects of iron chelators may rely in part on normalization of Lcn2 levels.

White matter T2 hyperintensities and blood-brain barrier disruption in the hyperacute stage of subarachnoid hemorrhage in male mice: The role of lipocalin-2.

AIMS: The current study examined whether white matter injury occurs in the hyperacute (4 hours) phase after subarachnoid hemorrhage (SAH) and the potential role of blood-brain barrier (BBB) disruption and an acute phase protein, lipocalin 2 (LCN2), in that injury. METHODS: Subarachnoid hemorrhage was induced by endovascular perforation in adult mice. First, wild-type (WT) mice underwent MRI 4 hours after SAH to detect white matter T2 hyperintensities. Second, changes in LCN2 expression and BBB disruption associated with the MRI findings were examined. Third, SAH-induced white matter injury at 4 hours was compared in WT and LCN2 knockout (LCN2 KO) mice. RESULTS: At 4 hours, most animals had uni- or bilateral white matter T2 hyperintensities after SAH in WT mice that were associated with BBB disruption and LCN2 upregulation. However, some disruption and LCN2 upregulation was also found in mice with no T2-hyperintensity lesion. In contrast, there were no white matter T2 hyperintensities in LCN2 KO mice after SAH. LCN2 deficiency also attenuated BBB disruption, myelin damage, and oligodendrocyte loss. CONCLUSIONS: Subarachnoid hemorrhage causes very early BBB disruption and LCN2 expression in white matter that is associated with and may precede T2 hyperintensities. LCN2 deletion attenuates MRI changes and pathological changes in white matter after SAH.

Interleukin-1ß Inhibition for Chronic Kidney Disease in Obese Mice With Type 2 Diabetes.

Inflammasome-driven release of interleukin(IL)-1ß is a central element of many forms of sterile inflammation and has been evident to promote the onset and progression of diabetic kidney disease. We microdissected glomerular and tubulointerstitial samples from kidney biopsies of patients with diabetic kidney disease and found expression of IL-1ß mRNA. Immunostaining of such kidney biopsies across a broad spectrum of diabetic kidney disease stages revealed IL-1ß positivity in a small subset of infiltrating immune cell. Thus, we speculated on a potential of IL-1ß as a therapeutic target and neutralizing the biological effects of murine IL-1ß with a novel monoclonal antibody in uninephrectomized diabetic db/db mice with progressive type 2 diabetes- and obesity-related single nephron hyperfiltration, podocyte loss, proteinuria, and progressive decline of total glomerular filtration rate (GFR). At 18 weeks albuminuric mice were randomized to intraperitoneal injections with either anti-IL-1ß or control IgG once weekly for 8 weeks. During this period, anti-IL-1ß IgG had no effect on food or fluid intake, body weight, and fasting glucose levels. At week 26, anti-IL-1ß IgG had reduced renal mRNA expression of kidney injury markers (Ngal) and fibrosis (Col1, a-Sma), significantly attenuated the progressive decline of GFR in hyperfiltrating diabetic mice, and preserved podocyte number without affecting albuminuria or indicators of single nephron hyperfiltration. No adverse effect were observed. Thus, IL-1ß contributes to the progression of chronic kidney disease in type 2 diabetes and might therefore be a valuable therapeutic target, potentially in combination with drugs with different mechanisms-of-action such as RAS and SGLT2 inhibitors.

G.I. pros: Antimicrobial defense in the gastrointestinal tract.

The gastrointestinal tract is a complex environment in which the host immune system interacts with a diverse array of microorganisms, both symbiotic and pathogenic. As such, mobilizing a rapid and appropriate antimicrobial response depending on the nature of each stimulus is crucial for maintaining the balance between homeostasis and inflammation in the gut. Here we focus on the mechanisms by which intestinal antimicrobial peptides regulate microbial communities during dysbiosis and infection. We also discuss classes of bacterial peptides that contribute to reducing enteric pathogen outgrowth. This review aims to provide a comprehensive overview on the interplay of diverse antimicrobial responses with enteric pathogens and the gut microbiota.

Expression and clinical correlation of NGAL and VEGF in endometrial carcinoma.

OBJECTIVE: To determine the expression of neutrophil gelatinase-associated lipocalin (NGAL) and vascular endothelial growth factor (VEGF) in endometrial carcinoma. PATIENTS AND METHODS: The clinical features (FIGO staging, pathological type, differentiation level, myometrial invasion depth and tumor size) and immunostaining analysis of endometrial carcinoma (30 cases), atypical hyperplasia (30 cases) and proliferative endometrial tissues (30 cases) were analyzed. Immunohistochemistry and reverse transcription PCR (RT-PCR) were performed to detect the expression of NGAL and VEGF. RESULTS: The positive tissue immunostaining and mRNA expression of NGAL and VEGF in endometrial carcinoma were significantly higher than in either atypical hyperplasia or proliferative endometrial tissues (p<0.05). The relative expression level of NGAL and VEGF was positively correlated with worse FIGO staging, higher differentiation level and a greater myometrial invasion depth (p<0.05); but not with patient age, pathological type or tumor size (p>0.05). CONCLUSIONS: The abnormal high expression of NGAL and VEGF observed in the endometrial carcinoma may be an important biomarker for early tumor diagnosis or as a novel target for therapeutic intervention.

Reduced Lipocalin 2 Expression Contributes to Vincristine Resistance in Human Colon Cancer Cells.

BACKGROUND: Vincristine (VCR) resistance can lead to cancer chemotherapy failure. Although changes in gene expression are responsible for drug resistance, the specific identities and roles of these genes remain unclear. OBJECTIVE: In this study, we aimed to identify differentially expressed genes and mechanisms of VCR resistance in colorectal cancer (CRC) cells. METHODS: A VCR-resistant CRC cell line (HCT-8/VCR) was established, and differentially expressed proteins between HCT-8 and HCT-8/VCR cells were screened using a human cytokine array; the results were confirmed by reverse transcription polymerase chain reaction and Western blotting. Furthermore, differentially expressed proteins were downregulated using siRNA, and cell proliferation and apoptosis were assessed by Cell Counting Kit-8 assay and flow cytometry, respectively. RESULTS: Compared with HCT-8 CRC cells, HCT-8/VCR cells showed downregulation of lipocalin 2 (LCN2). We found that siRNA-mediated downregulation of LCN2 in HCT-8 cells significantly increased VCR resistance. Furthermore, when we downregulated LCN2, we observed significant decreases in apoptosis, but no significant effect on cell cycle. CONCLUSION: Overall, these results demonstrate that LCN2 plays an important role in VCR resistance and is a potential therapeutic target for this disease.

Experimental and Human Evidence for Lipocalin-2 (Neutrophil Gelatinase-Associated Lipocalin [NGAL]) in the Development of Cardiac Hypertrophy and heart failure.

BACKGROUND: Cardiac hypertrophy increases the risk of developing heart failure and cardiovascular death. The neutrophil inflammatory protein, lipocalin-2 (LCN2/NGAL), is elevated in certain forms of cardiac hypertrophy and acute heart failure. However, a specific role for LCN2 in predisposition and etiology of hypertrophy and the relevant genetic determinants are unclear. Here, we defined the role of LCN2 in concentric cardiac hypertrophy in terms of pathophysiology, inflammatory expression networks, and genomic determinants. METHODS AND RESULTS: We used 3 experimental models: a polygenic model of cardiac hypertrophy and heart failure, a model of intrauterine growth restriction and Lcn2-knockout mouse; cultured cardiomyocytes; and 2 human cohorts: 114 type 2 diabetes mellitus patients and 2064 healthy subjects of the YFS (Young Finns Study). In hypertrophic heart rats, cardiac and circulating Lcn2 was significantly overexpressed before, during, and after development of cardiac hypertrophy and heart failure. Lcn2 expression was increased in hypertrophic hearts in a model of intrauterine growth restriction, whereas Lcn2-knockout mice had smaller hearts. In cultured cardiomyocytes, Lcn2 activated molecular hypertrophic pathways and increased cell size, but reduced proliferation and cell numbers. Increased LCN2 was associated with cardiac hypertrophy and diastolic dysfunction in diabetes mellitus. In the YFS, LCN2 expression was associated with body mass index and cardiac mass and with levels of inflammatory markers. The single-nucleotide polymorphism, rs13297295, located near LCN2 defined a significant cis-eQTL for LCN2 expression. CONCLUSIONS: Direct effects of LCN2 on cardiomyocyte size and number and the consistent associations in experimental and human analyses reveal a central role for LCN2 in the ontogeny of cardiac hypertrophy and heart failure.

IL-36R signalling activates intestinal epithelial cells and fibroblasts and promotes mucosal healing in vivo.

OBJECTIVE: Interleukin (IL)-36R signalling plays a proinflammatory role in different organs including the skin, but the expression of IL-36R ligands and their molecular function in intestinal inflammation are largely unknown. DESIGN: We studied the characteristics of IL-36R ligand expression in IBDs and experimental colitis. The functional role of IL-36R signalling in the intestine was addressed in experimental colitis and wound healing models in vivo by using mice with defective IL-36R signalling (IL-36R-/-) or Myd88, neutralising anti-IL-36R antibodies, recombinant IL-36R ligands and RNA-seq genome expression analysis. RESULTS: Expression of IL-36α and IL-36γ was significantly elevated in active human IBD and experimental colitis. While IL-36γ was predominantly detected in nuclei of the intestinal epithelium, IL-36α was mainly found in the cytoplasm of CD14+ inflammatory macrophages. Functional studies showed that defective IL-36R signalling causes high susceptibility to acute dextran sodium sulfate colitis and impairs wound healing. Mechanistically, IL-36R ligands released upon mucosal damage activated IL-36R+ colonic fibroblasts via Myd88 thereby inducing expression of chemokines, granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-6. Moreover, they induced proliferation of intestinal epithelial cells (IECs) and expression of the antimicrobial protein lipocalin 2. Finally, treatment of experimental intestinal wounds with IL-36R ligands significantly accelerated mucosal healing in vivo. CONCLUSIONS: IL-36R signalling is activated upon intestinal damage, stimulates IECs and fibroblasts and drives mucosal healing. Modulation of the IL-36R pathway emerges as a potential therapeutic strategy for induction of mucosal healing in IBD.

Chronic psychological stress and high-fat high-fructose diet disrupt metabolic and inflammatory gene networks in the brain, liver, and gut and promote behavioral deficits in mice.

The mechanisms underlying the association between chronic psychological stress, development of metabolic syndrome (MetS), and behavioral impairment in obesity are poorly understood. The aim of the present study was to assess the effects of mild chronic psychological stress on metabolic, inflammatory, and behavioral profiles in a mouse model of diet-induced obesity. We hypothesized that (1) high-fat high-fructose diet (HFHF) and psychological stress would synergize to mediate the impact of inflammation on the central nervous system in the presence of behavioral dysfunction, and that (2) HFHF and stress interactions would impact insulin and lipid metabolism. C57Bl/6 male mice underwent a combination of HFHF and two weeks of chronic psychological stress. MetS-related conditions were assessed using untargeted plasma metabolomics, and structural and immune changes in the gut and liver were evaluated. Inflammation was measured in plasma, liver, gut, and brain. Our results show a complex interplay of diet and stress on gut alterations, energetic homeostasis, lipid metabolism, and plasma insulin levels. Psychological stress and HFHF diet promoted changes in intestinal tight junctions proteins and increases in insulin resistance and plasma cholesterol, and impacted the RNA expression of inflammatory factors in the hippocampus. Stress promoted an adaptive anti-inflammatory profile in the hippocampus that was abolished by diet treatment. HFHF increased hippocampal and hepatic Lcn2 mRNA expression as well as LCN2 plasma levels. Behavioral changes were associated with HFHF and stress. Collectively, these results suggest that diet and stress as pervasive factors exacerbate MetS-related conditions through an inflammatory mechanism that ultimately can impact behavior. This rodent model may prove useful for identification of possible biomarkers and therapeutic targets to treat metabolic syndrome and mood disorders.

Expression of Neutrophil Gelatinase-Associated Lipocalin and Kidney Injury Molecule-1 in Wilms Tumor.

OBJECTIVE: Neutrophil gelatinase-associated lipocalin (NGAL) and Kidney injury molecule-1 (KIM-1) play important roles in both immunity and cell proliferation. It was reported previously that they are overexpressed in various human cancers. The present study was undertaken to examine the expressions of NGAL and KIM-1 in Wilms Tumors. MATERIAL AND METHOD: Tissue samples of 50 Wilms Tumors were evaluated and underwent immunhistochemical staining for NGAL and KIM-1 protein expressions. The correlations between them, and some clinical prognostic factors such as tumor weight, stage and histological features were also evaluated. RESULTS: Twenty-three (46%) of the cases were male while 27 (54%) were female. The mean age was found to be 3.26±2 years. The average tumor size was 9.16 ± 2.9 cm in diameter and the average weight of the kidney was 478±312 gr. Thirteen (26%) cases were stage I, 18 (36%) cases were stage II, 7 (14%) cases were stage III, and 6 (12%) cases were stage IV. Thirty-nine cases were alive (78%), while 11 cases (22%) were deceased. Mean overall survival time was 68.2±39.5 (2-148) months. NGAL expression was negative in all tumors except the neutrophils within the tumors. KIM-1 expression was positive in 37 tumors (74%), while it was absent in 13 tumors (26%). Using Mann-Whitney U Analysis, KIM-1 expression was found to be associated with the stage of the tumor (p=0.027). CONCLUSION: The preliminary data indicates that KIM-1 expression may be associated with stage in Wilms Tumor. However, further studies are needed to validate these pilot observations and to clarify the functional and mechanistic significance of this relevance.

The Nuclear Protein IκBζ Forms a Transcriptionally Active Complex with Nuclear Factor-κB (NF-κB) p50 and the Lcn2 Promoter via the N- and C-terminal Ankyrin Repeat Motifs.

The nuclear protein IκBζ, comprising the N-terminal trans-activation domain and the C-terminal ankyrin repeat (ANK) domain composed of seven ANK motifs, activates transcription of a subset of nuclear factor-κB (NF-κB)-dependent innate immune genes such as Lcn2 encoding the antibacterial protein lipocalin-2. Lcn2 activation requires formation of a complex containing IκBζ and NF-κB p50, a transcription factor that harbors the DNA-binding Rel homology region but lacks a trans-activation domain, on the promoter with the canonical NF-κB-binding site (κB site) and its downstream cytosine-rich element. Here we show that IκBζ productively interacts with p50 via Asp-451 in the N terminus of ANK1, a residue that is evolutionarily conserved among IκBζ and the related nuclear IκB proteins Bcl-3 and IκBNS Threonine substitution for Asp-451 abrogates direct association with the κB-site-binding protein p50, complex formation with the Lcn2 promoter DNA, and activation of Lcn2 transcription. The basic residues Lys-717 and Lys-719 in the C-terminal region of ANK7 contribute to IκBζ binding to the Lcn2 promoter, probably via interaction with the cytosine-rich element required for Lcn2 activation; glutamate substitution for both lysines results in a loss of transcriptionally active complex formation without affecting direct contact of IκBζ with p50. Both termini of the ANK domain in Bcl-3 and IκBNS function in a manner similar to that of IκBζ to interact with promoter DNA, indicating a common mechanism in which the nuclear IκBs form a regulatory complex with NF-κB and promoter DNA via the invariant aspartate in ANK1 and the conserved basic residues in ANK7.

Progesterone induces the expression of lipocalin-2 through Akt-c-Myc pathway during mouse decidualization.

Lipocalin-2 (Lcn2) is a small glycoprotein involved in a number of biological processes such as inflammation and antibacterial response. In our study, Lcn2 is expressed in the subluminal stromal cells at implantation site on day 5 of pregnancy. The expression of Lcn2 in stromal cells is under the control of progesterone through Akt-c-Myc signaling pathway. Data from Lcn2 knockdown and recombinant protein treatments indicate that Lcn2 promotes mPGES-1 expression in stromal cells. The expression of Lcn2 and mPGES-1 is strongly stimulated by lipopolysaccharide (LPS), indicating that Lcn2 mediates LPS-induced inflammation. These findings shed light on the role of Lcn2 during decidualization.

Acute Stress Responses Are Early Molecular Events of Retinal Degeneration in Abca4-/-Rdh8-/- Mice After Light Exposure.

PURPOSE: Mice lacking ATP-binding cassette transporter 4 (ABCA4) and retinol dehydrogenase 8 (RDH8) mimic features of human Stargardt disease and age-related macular degeneration. RNA-sequencing of whole eyes was done to study early gene expression changes in Abca4-/-Rdh8-/- mice. METHODS: Abca4-/-Rdh8-/- mice at 4 weeks of age were exposed to intense light. Total RNA was extracted from whole eyes and used to generate RNA libraries that were paired-end sequenced on the Illumina HiSeq 2500 device. Differentially expressed genes were annotated using Gene set enrichment analysis (GSEA). Selected genes in enriched pathways exhibiting differential expression were validated using quantitative qRT-PCR and ELISA. RESULTS: Transcriptome analysis of the whole eye identified 200 genes that were differentially expressed 24 hours after light exposure compared to no light in Abca4-/-Rdh8-/- mice. Expression of several visual cycle and photoreceptor genes were decreased, indicative of photoreceptor/RPE cell death. Gene categories of early stress response genes, inflammatory cytokines, immune factors, and JAK STAT components were upregulated. Lipocalin 2 (Lcn2) was the most upregulated early stress response gene identified. Protein LCN2 was produced by RPE cells and the neural retina after intense light exposure as well as in cultured RPE cells from mice and humans incubated with lipopolysaccharide or photoreceptor outer segments. CONCLUSIONS: Identification of important mediators involved in the crosstalk between the acute stress response and immune activation in RPE cells and the neural retina, such as LCN2, provide novel molecular targets for reducing cellular stress during retinal degeneration.

Evidence for participation of neutrophil gelatinase-associated lipocalin/matrix metalloproteinase-9 (NGAL•MMP-9) complex in the inflammatory response to infection in pregnancies complicated by preterm birth.

PROBLEM: Neutrophil gelatinase-associated lipocalin (NGAL) is expressed in neutrophils and involved in innate immunity by sequestering iron. NGAL's ability to complex with matrix metalloproteinase-9 (MMP-9) and extend its gelatinolytic activity led us to investigate its role in pregnancies complicated by preterm birth (PTB) and intra-amniotic infection/inflammation (IAI). METHOD OF STUDY: We assayed the amniotic fluid (AF) levels of NGAL and MMP-9 in 308 women that had a clinically indicated amniocentesis and a normal pregnancy outcome or PTB. qRT-PCR was employed to determine NGAL mRNA expression of placental villous trophoblast and amniochorion. Immunohistochemistry was used for cellular localization. RESULTS: AF NGAL levels were gestational age-regulated. Women with IAI and PTB had significantly higher levels of NGAL, MMP-9 and NGAL•MMP-9 complex. CONCLUSION: The amniochorion is a source of NGAL and similarly to other inflammatory conditions, this protein may augment the collagenolytic effect of MMP-9 and modulate host-microbe interactions in pregnancies complicated by IAI.

Lipocalin 2 prevents oral cancer metastasis through carbonic anhydrase IX inhibition and is associated with favourable prognosis.

Lipocalin 2 (LCN2), a secreted glycoprotein, is up- or downregulated in different human cancers. At present, the functional role of LCN2 in the progression of oral squamous cell carcinoma (OSCC), which accounts for most head and neck cancers, remains poorly understood, particularly with respect to its involvement in invasion and metastasis. In this study, we observed that LCN2 expression decreased in patients with OSCC and lymph node metastasis compared with that in patients without metastasis. A higher LCN2 expression correlated with the survival of patients with OSCC. Furthermore, LCN2 overexpression in OSCC cells reduced in vitro migration and invasion and in vivo metastasis, whereas its silencing induced an increase in cell motility. Mechanistically, LCN2 inhibited the cell motility of OSCC cells through hypoxia-inducible factor (HIF)-1α-dependent transcriptional inhibition of the carbonic anhydrase IX (CAIX). CAIX overexpression relieved the migration inhibition imposed by LCN2 overexpression in OSCC cells. Moreover, a microRNA (miR) analysis revealed that LCN2 can suppress CAIX expression and cell migration through miR-4505 induction. Examination of tumour tissues from patients with OSCC and OSCC-transplanted mice revealed an inverse correlation between LCN2 and CAIX expression. Furthermore, patients with LCN2(strong)/CAIX(weak) revealed the lowest frequency of lymph node metastasis and the longest survival. Our findings suggest that LCN2 suppresses tumour metastasis by targeting the transcriptional and post-transcriptional regulation of CAIX in OSCC cells. LCN2 overexpression may be a novel OSCC treatment strategy and a useful biomarker for predicting OSCC progression.