Gene Expression In Immune System Research Articles

Characteristics of RNA Stabilizer RNApro for Peripheral Blood Collection.

Peripheral blood is the most practical tissue for human immune system gene expression profiling because it is easily accessible, whereas the site of primary infection in certain diseases may not be easily accessible. Due to the ex vivo instability of RNA transcripts, a key challenge in the gene expression analysis of blood samples is the rapid sample handling and stabilization of the mRNA by adding an RNA preservative (PAXgeneTM Blood RNA Tubes, TempusTM Blood RNA tubes, RNAlater Stabilization Reagent, RNAgard® Blood Tubes). BioMole (Turin, Italy) has developed a novel blood stabilizer, called RNApro, in which RNA is stabilized during phlebotomy and sample storage. In this study, RNApro performance intended as RNA yield, integrity, and stability was evaluated. Our results show that blood samples stored at -80 °C and re-extracted after 7 years show no differences in terms of quantity, quality, and amplificability. The samples in the RNAlater stabilization solution can be stored at room temperature for up to one week or at 4 °C for up to one month. Similar results can also be observed for PAXgene tubes, Tempus tubes, and RNAgard tubes. In agreement with these data, the RNApro stabilization solution preserves the RNA from degradation for up to 1 month at 4 °C and 1 week at room temperature. RNApro can be stored indifferently at -80, -20, 4 °C, or room temperature for up to 2 months after, and then could be stored at -80 °C for up to seven years. In summary, our study is the first to analyze the performance of an RNA stabilizer called RNApro. We can conclude that several studies have shown significant differences in gene expression analysis when the sample was preserved in different RNA stabilizers. Therefore, it is desirable to standardize the method of nucleic acid conservation when comparing data from transcriptomic analyses.

APOBEC3G Is a p53-Dependent Restriction Factor in Respiratory Syncytial Virus Infection of Human Cells Included in the p53/Immune Axis.

Identifying and understanding genetic factors that influence the propagation of the human respiratory syncytial virus (RSV) can lead to health benefits and possibly augment recent vaccine approaches. We previously identified a p53/immune axis in which the tumor suppressor p53 directly regulates the expression of immune system genes, including the seven members of the APOBEC3 family of DNA cytidine deaminases (A3), which are innate immune sentinels against viral infections. Here, we examined the potential p53 and A3 influence in RSV infection, as well as the overall p53-dependent cellular and p53/immune axis responses to infection. Using a paired p53 model system of p53+ and p53- human lung tumor cells, we found that RSV infection activates p53, leading to the altered p53-dependent expression of A3D, A3F, and A3G, along with p53 site-specific binding. Focusing on A3G because of its 10-fold-greater p53 responsiveness to RSV, the overexpression of A3G can reduce RSV viral replication and syncytial formation. We also observed that RSV-infected cells undergo p53-dependent apoptosis. The study was expanded to globally address at the transcriptional level the p53/immune axis response to RSV. Nearly 100 genes can be directly targeted by the p53/immune axis during RSV infection based on our p53BAER analysis (Binding And Expression Resource). Overall, we identify A3G as a potential p53-responsive restriction factor in RSV infection. These findings have significant implications for RSV clinical and therapeutic studies and other p53-influenced viral infections, including using p53 adjuvants to boost the response of A3 genes.

Effects of Dietary Sodium Propionate on Growth, Digestive Enzyme Activity, and Expression of Immune System Genes in Juveniles of Tropical Gar (Atractosteus tropicus)

We determined the effects of sodium propionate (SP) added to the diets of Atractosteus tropicus juveniles with respect to the growth, survival, digestive enzyme activity, and expression of genes that are associated with the immune system. Five treatments (0, 0.5, 1.0, 1.5, and 2.0%) were evaluated in triplicate on 180 fish (3.65 ± 0.12 g) distributed among 15 (70 L) tanks. The juveniles were fed five times a day with 5% feed in relation to the biomass of the organism. The treatment with 0.5% SP showed a final weight value of 25.7 ± 4.5 g, absolute weight of gain (AWG) of 21.93 ± 4.39 g, and specific growth rate (SGR) of 3.1 ± 0.26. Treatments with 1.5 and 2.0% SP showed the highest survival (91.6%). The control group (0%) showed a greater activity of lipases. There was a tendency that the highest activity of alkaline proteases and chymotrypsin occurred in the 0 and 0.5% treatments. The maximum relative expression of the genes ocln, muc2, and nod2 occurred in the 1.5% treatment. The inclusion of SP in the diet of A. tropicus juveniles could benefit the activity of some digestive enzymes as well as the expression of genes related to the function of the intestinal barrier, therefore benefitting the survival of the organisms.

Positive and negative emotion are associated with generalized transcriptional activation in immune cells

Alterations in immune system gene expression have been implicated in psychopathology, but it remains unclear whether similar associations occur for intraindividual variations in emotion. The present study examined whether positive emotion and negative emotion were related to expression of pro-inflammatory and antiviral genes in circulating leukocytes from a community sample of 90 adolescents (Mage = 16.3 years, SD = 0.7; 51.1% female). Adolescents reported their positive emotion and negative emotion and provided blood samples twice, five weeks apart. Using a multilevel analytic framework, we found that within-individual increases in positive emotion were associated with reduced expression of both pro-inflammatory and Type I interferon (IFN) response genes, even after adjusting for demographic and biological covariates, and for leukocyte subset abundance. By contrast, increases in negative emotion were related to higher expression of pro-inflammatory and Type I IFN genes. When tested in the same model, only associations with positive emotion emerged as significant, and increases in overall emotional valence were associated with both lower pro-inflammatory and antiviral gene expression. These results are distinct from the previously observed Conserved Transcriptional Response to Adversity (CTRA) gene regulation pattern characterized by reciprocal changes in pro-inflammatory and antiviral gene expression and may reflect alterations in generalized immunologic activation. These findings highlight one biological pathway by which emotion may potentially impact health and physiological function in the context of the immune system, and future studies can investigate whether fostering positive emotion may promote adolescent health through changes in the immune system.

Colony Transport Affects the Expression of Some Genes Related to the Apis mellifera L. Immune System

Migratory beekeeping can harm the bee colonies if not executed properly. Here, colonies of Apis mellifera were transported (for one or two hours) or not, following proper technical standards. To analyze gene expression (defensin-1, abaecin, and HSP70), forager bees were collected immediately, 24, and 72 hours after transportation. Bee mortality and population growth were measured before and after transportation. This study concludes that transporting honey bee colonies for 2 hours promotes immune system gene expression, although there are no significant changes in bee mortality and population growth of the colonies.

Microplastic-Contaminated Feed Interferes with Antioxidant Enzyme and Lysozyme Gene Expression of Pacific White Shrimp (Litopenaeus vannamei) Leading to Hepatopancreas Damage and Increased Mortality

Microplastic pollution can interfere with aquatic animal health and nonspecific immunity, increasing the potential for pathogen infection in crustaceans. However, the long-term effects of microplastics on crustacean immunity are less understood, especially regarding their toxicity in Pacific white shrimp (Litopenaeus vannamei). Effects of high-density polyethylene microplastics (HDPE-MPs) in feed on the mortality rate, hepatopancreas, and nonspecific immune system gene expression of Pacific white shrimp are presented. The LC50 at day 28 of HDPE-MP exposure was determined as 3.074% HDPE-MP in feed. A significant upregulation of the superoxide dismutase (SOD) and glutathione peroxidase (GPx) genes was observed in shrimp that were fed with 0.1 and 0.5% of HDPE-MP; then, they were downregulated significantly, except for the SOD gene expression of shrimp fed with 0.1% of HDPE-MP. The lysozyme (LYZ) gene was upregulated significantly in shrimp that were fed with 0.5, 1, and 3% HDPE-MP for 7 days and downregulated significantly in HDPE-receiving groups for at least 14 days. Significant histopathological changes in the hepatopancreas were observed in the treatment groups. The histopathological score of each lesion was correlated with the increase in HDPE-MP concentration. This study shows that the ingestion of HDPE microplastics can alter the expression of nonspecific immune system genes and damage the hepatopancreas in Pacific white shrimp.

Metformin attenuates depressive-like behaviour of methamphetamine withdrawal in mice: A mechanistic approach

Objectives Methamphetamine (METH) as a potent psychostimulant drug with a high potency of dependence rate that results in neurotoxicity has become a major drug of abuse in many parts of the world. Unfortunately, there is limited evidence regarding treatment of METH withdrawal syndrome. Therefore, we aimed to investigate whether metformin mitigate the methamphetamine (METH) withdrawal syndrome in male mice. Based on the literature, depression and anxiety are the major METH withdrawal symptoms. Methods Here, METH (2 mg/kg) was administered to mice twice a day for 14 constitutive days to induce animal model of METH-induced withdrawal syndrome. To do this, mice in control group and those with METH withdrawal syndrome were divided into treatment (receiving metformin in 3 doses of 50, 100 and 200 mg/kg for 10 days) and non-treatment sub-groups. Following the behavioural test, the animals were sacrificed; their hippocampus was dissected to measure oxidative stress parameters and expression of cellular energy homeostasis and immune-inflammatory genes. Results Our data revealed that metformin provoked antidepressant effects in behavioural tests through AMPK overexpression as an important mitochondrial energetic sensor and inhibition of Tlr4 overexpression in the immune system gene expression. In addition, metformin was able to improve oxidative stress biomarkers and neuronal damage in the hippocampus and restore cellular energy homeostasis and immune system gene expression. Conclusions The data suggested that metformin can influence the hippocampus through targeting mitochondria and their performance, and consequently, neuroinflammation responses and brain metabolic changes. It is supposed to be a new therapeutic option in clinical trials of depression and anxiety following METH withdrawal treatment.

MO1005: Analysis of Expression of Immune System Genes that are Responsible for Activation and Inhibition of T-Cell Immune Response in Renal Transplant Recipients after Extracorporeal Photochemotherapy

Abstract BACKGROUND AND AIMS Due to the increasing need for kidney transplantation, there is a demand for new methods of preventing rejection of the transplant based on the mechanisms of natural regulation of the immune response. Extracorporeal photopheresis (ECP) is one of these technologies whose clinical efficacy in the prevention and treatment of acute renal transplant rejection has been confirmed by many studies, but the mechanism of its effect has not been fully understood. Characterization of the genes responsible for the activation and inhibition of the immune response during ECP is an important step towards the modification of immunosuppressive therapy. The aim of this study was to analyze the changes in the expression level of the genes responsible for the activation and inhibition of the T-cell response in the recipients of the renal graft after ECP. METHOD An open cohort randomized trial was conducted with 20 patients who underwent a single group cadaver kidney transplantation from an unrelated donor. The recipients of the compared groups received standard immunosuppressive therapy and the same therapy in combination with ECP procedures, according to the study protocol. Each recipient of the main group received 15 sessions of ECP within 6 months. The level of gene expression was analyzed by real-time RT-PCR. Comparison of the study groups was carried out at point 1 (1st–4th day) and 2 (30th day). RESULTS The analysis of clinical data did not show statistically significant clinical differences, but in the specific consideration of each pair of cases, the tendency to some improvement in the transplant organ engraftment rate in the early terms after the transplant, and better survival of the graft and the recipient was revealed. For instance, patients of the main group revealed no transplant rejection by clinical laboratory signs as well as the results of histological examination of biopsy specimens. In the comparison group, acute graft rejection was clinically diagnosed and was confirmed by histological examination as humoral type rejection. The level of expression of genes involved in immune response regulation (CD28, CTLA4, PDL1 and FOXP3) and cytokine genes (TNFA, IL1, IL2, IL10 and IFNG) in the studied groups was determined. A decrease in the level of expression of the genes for the proinflammatory cytokines IL1, IL2 and IFNG was noted by the 30th day after transplantation. At the same time, the expression of the genes PDL1 and FOXP3, which regulate the development of the regulatory response, significantly increased by the 30th day after transplantation. Correlations of the level of gene expression with the presence/absence of rejection episodes have not been established. To assess the effect of the ECP on these gene expression, a comparison was made of the level sof transcript representation of patients receiving standard immunotherapy and immunotherapy in combination with ECP, and a significant increase in the expression of PDL1 and FOXP3 genes (Fig. 1A and B) was found in the group of patients who underwent ECP procedures (P = .0009 and .0013, respectively). CONCLUSION According to the results of comparative analysis, the level of expression of pro-inflammatory cytokines' genes IL1, IL2 and IFNG decreased to the 30th day after transplantation. A 30-day comparison between groups shows an increase in the expression level of the PDL1 and FOXP3 genes. Increased expression of these genes may be associated with the activation of regulatory T-cells and the development of peripheral tolerance and can become a criterion for the modification of immunosuppressive therapy.

Are owned dogs or stray dogs more prepared to diseases? A comparative study of immune system gene expression of perforin and granzymes.

Stray dogs are inevitably exposed to more infections than owned dogs living indoor. However, no studies have investigated whether the immune system gene expression differs between owned dogs kept in better care conditions and stray dogs living outside. To investigate this, blood samples were taken from 90 dogs (45 owned and 45 stray dogs) that were checked and confirmed as healthy. By using qPCR, the amples were analyzed for the expression of the perforin, granzyme A and granzyme B genes, which are associated with the activation of apoptotic pathways in the immune system. We found that the perforin and granzyme A gene expression levels were higher in stray dogs although the differences were not statistically significant. On the other hand, a 2.81 times higher and a notable difference (P < 0.001) was found in the expression level of granzyme B gene in stray dogs. These results indicate that the immune system in stray dogs might be more prepared for diseases than that of the owned dogs and the granzyme B gene plays a more dominant role in the immune response than granzyme A and perforin.

Differences in peripheral immune system gene expression in frontotemporal degeneration.

The peripheral immune system has a key pathophysiologic role in Frontotemporal degeneration (FTD). We sought a comprehensive transcriptome-wide evaluation of gene expression alterations unique to the peripheral immune system in FTD compared to healthy controls and amyotrophic lateral sclerosis.Nineteen subjects with FTD with 19 matched healthy controls and 9 subjects with amyotrophic lateral sclerosis underwent isolation of peripheral blood mononuclear cells (PBMCs) which then underwent bulk ribonucleic acid sequencing.There was increased expression in genes associated with CD19+ B-cells, CD4+ T-cells, and CD8+ T-cells in FTD participants compared to healthy controls. In contrast, there was decreased expression in CD33+ myeloid cells, CD14+ monocytes, BDCA4+ dendritic cells, and CD56+ natural killer cells in FTD and healthy controls. Additionally, there was decreased expression is seen in associated with 2 molecular processes: autophagy with phagosomes and lysosomes, and protein processing/export. Significantly downregulated in PBMCs of FTD subjects were genes involved in antigen processing and presentation as well as lysosomal lumen formation compared to healthy control PBMCs.Our findings that the immune signature based on gene expression in PBMCs of FTD participants favors adaptive immune cells compared to innate immune cells. And decreased expression in genes associated with phagosomes and lysosomes in PBMCs of FTD participants compared to healthy controls.

Effects of Treatment with Lactobacilli on Necrotic Enteritis in Broiler Chickens.

Growth promoter antibiotics have been commonly used for the control of necrotic enteritis (NE) in broilers for decades. However, due to a ban on the use of these antibiotics, alternatives such as probiotics have been tested widely for NE control. The present study tested the efficacy of four different species of lactobacilli (two isolates of Lactobacillus johnsonii and one of Ligilactobacillus (L.) salivarius, Limosilactobacillus (L.) reuteri, and L. crispatus) against NE. Day-old male broiler chickens were divided into six groups and orally inoculated with 1 × 107 or 1 × 108 colony-forming units (CFU) of lactobacilli on 1, 7, 14, and 20days of age. While negative and positive control groups did not receive lactobacilli, the latter was challenged with Clostridium perfringens (CP). Chickens, at 21days old, were challenged for 3days with 3 × 108CFU of a virulent strain of CP. Tissues were collected for immune system gene expression, immunophenotyping, intestinal histomorphometry, and microbiota analysis. Lactobacilli inoculation conferred partial protection in chickens against NE, marked by lowered lesion scores and improved villus:crypt ratio. Immunomodulatory effects were demonstrated by the significant alteration of interferon (IFN)-γ, interleukin (IL)-1β, IL-2, IL-12p35, IL-17, and transforming growth factor beta (TGF-β) gene transcription in the duodenum and jejunum as well as subtle changes in the frequency of CD8 + T cells and B cells in the cecal tonsil of the treated chickens. Microbiota analysis showed increased levels of some bacterial phyla including Actinobacteria, Lactobacillaceae, and Firmicutes. In conclusion, these findings suggest that the use of certain lactobacilli can reduce NE severity and modulate immune responses and intestinal microbiota composition in chickens.

Supering Apis mellifera (Hymenoptera, Apidae) beehives impairs honey production and biomarker genes

For honey production, beekeepers add one or more supers to the hives to allow honeybees to store their products. However, the increase in hive space can affect the social and health organization in the colony, promoting stress. This study assessed the management of honey production, physicochemical honey properties, population development, and forages immune system gene expression patterns to be used as biomarker for monitoring beekeeping welfare. The treatments comprised 40 beehives divided in four treatments. Treatment 1 - control, supers added according to storage necessity. Treatments 2, 3, and 4 presented two, three, and four supers at the beginning of the experiment, respectively. T1 presented greater honey production (39.4 % increased). No difference in open brood area in the colonies was observed and honey properties and only T2 showed closed brood area higher than the other treatments. Foragers from T4 showed higher catalase and defensin gene expression at the middle-end experiment. Thus, the increasing internal space at the beginning of honey season can affect honey production and immune system of foragers. Catalase and defensin can be used as biomarkers for monitoring honey production welfare.

Neuroprotective Effect of Apigenin on Depressive-Like Behavior: Mechanistic Approach.

Apigenin, as a natural flavonoid present in several plants is characterized with potential anticancer, antioxidant, and anti-inflammatory properties. Recent studies proposed that apigenin affects depression disorder through unknown mechanistic pathways. The effects of apigenin's anti-depressive properties on streptozocin-mediated depression have been investigated through the evaluation of behavioral tests, oxidative stress, cellular energy homeostasis and inflammatory responses. The results demonstrated anti-depressive properties of apigenin in behavioral test including forced swimming and splash tests and oxidative stress biomarkers such as reduced glutathione, lipid peroxidation, total antioxidant power and coenzyme Q10 levels. Apigenin, also, demonstrated its regulatory potency in cellular energy homeostasis and immune system gene expression through inhibiting Nlrp3 and Tlr4 overexpression. Furthermore, failure in energy production as the key factor in various psychiatric disorders was reversed by apigenin modulating effect on AMPK gene expression. Overall, 20mg/kg of apigenin was recognized as the dose suitable for minimizing the undesirable adverse effects in the STZ-mediated depression model proposed in this study. Our data suggested that apigenin could be able to adjust behavioral dysfunction, biochemical biomarkers and recovered cellular antioxidant level in depressed animals. The surprising results were achieved by raise in COQ10 level, which could regulate the overexpression of the AMPK gene in stressful conditions. The regulatory effect of apigenin in inflammatory signaling pathways such as Nlrp3, and Tlr4 gene expression was studied at the surface part of the hippocampus.

Abstract PO022: Somatic mutations in tumor infiltrating lymphocytes can affect the expression of immune system genes in the tumor microenvironment

Abstract Avoiding immune destruction is an emerging hallmark of cancer. It includes tumor cell mutations and epigenetic modifications that allow tumor cells to either evade detection by the immune system, or render the immune response ineffective. However, mutations in tumor infiltrating lymphocytes that inhibit immune response to tumor growth, have not been specifically identified as a hallmark of cancer. We have identified four distinct somatic mutations in tumor infiltrating lymphocytes that affect the expression of immune system genes in tumor samples, suggesting that these lymphocytic mutation may cause a deficient immune response, contributing to cancer progression. These rare mutations occurred frequently in specific cancer types. The T292Nfs*18 mutation in the FETUB gene, R216Gfs*20 in CCDC43, N377D in CLEC4M and V78Gfs*46 in RAB11FIP4 occurred in 13% of breast invasive carcinoma, 41% of colon adenocarcinoma, 12% of ovarian cystadenocarcinoma (OV) and 16% of OV samples respectively, compared to 0.002%, 0.003%, 0.014% and 0.001% in aggregated variant databases. These variants were found in 7-20% of DNA sequences in both tumor and matched normal blood samples, indicating that they originated in tumor infiltrating lymphocytes, rather than in the tumor cells. We argue that these mutations are unlikely to be passenger mutations since each of them occur in only 0-2% of normal tissue samples and in less than 2% of tumor samples from 24-32 of 32 cancer types other than the one noted above for each of the four mutations. The set of differentially expressed genes (FDR&amp;lt;0.05) in samples with each of the four variants, compared to samples without the variant, included 22-445 genes associated with the immune system pathway, suggesting a possible mechanism by which these mutations may affect tumor growth. Given the myriad of diverse mechanisms and genes implicated in cancer, it is not unexpected that mutations in tumor infiltrating lymphocytes could contribute to cancer progression. With further validation, genetic lesions in tumor infiltrating lymphocytes may represent a previously unstudied mechanism affecting tumor growth, and a new class of therapeutic targets. These mutations may also represent potential “somatic” risk markers that may present with age, supplementing traditional germline risk markers. Citation Format: Ramu Anandakrishnan, Robin T. Varghese, Nicholas A. Kinney, Harold R. Garner. Somatic mutations in tumor infiltrating lymphocytes can affect the expression of immune system genes in the tumor microenvironment [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2020 Oct 19-20. Philadelphia (PA): AACR; Cancer Immunol Res 2021;9(2 Suppl):Abstract nr PO022.

Анализ экспрессии генов иммунной системы, ответственных за активацию и ингибирование Т-клеточного иммунного ответа, у реципиентов почечного трансплантата после ЭКФ

Анализ экспрессии генов иммунной системы, ответственных за активацию и ингибирование Т-клеточного иммунного ответа, у реципиентов почечного трансплантата после ЭКФ

Microplastics induce transcriptional changes, immune response and behavioral alterations in adult zebrafish

Microplastics have become pervasive environmental pollutants in both freshwater and marine ecosystems. The presence of microplastics have been recorded in the tissues of many wild fish species, and laboratory studies have demonstrated that microplastics can exert adverse health effects. To further investigate the biological mechanisms underlying microplastics toxicity we applied an integrated approach, analyzing the effects of microplastics at transcriptomic, histological and behavioral level. Adult zebrafish have been exposed to two concentrations of high-density polyethylene and polystyrene microplastics for twenty days. Transcriptomic results indicate alterations in the expression of immune system genes and the down-regulation of genes correlated with epithelium integrity and lipid metabolism. The transcriptomic findings are supported by tissue alterations and higher occurrence of neutrophils observed in gills and intestinal epithelium. Even the daily rhythm of activity of zebrafish appears to be affected, although the regular pattern of activity is recovered over time. Considering the transcriptomic and histological findings reported, we hypothesize that the effects on mucosal epithelium integrity and immune response could potentially reduce the organism defense against pathogens, and lead to a different utilization of energy stores.

Evidence of a hydraulically challenging reach serving as a barrier for the upstream migration of infection-burdened adult steelhead.

Anadromous fishes such as steelhead trout, Oncorhynchus mykiss, are exposed to a suite of infectious agents and migratory challenges during their freshwater migrations. We assessed infectious agent load and richness and immune system gene expression in gill tissue of Bulkley River (British Columbia, CA) steelhead captured at and upstream of a migratory barrier to evaluate whether infectious burdens impacted migration success. We further considered the potential influences of water temperature, sex and fish size on host infectious agents and transcription profiles. There were eight infectious agents detected in steelhead gill tissue, with high prevalence of the bacteria Candidatus Branchiomonas cysticola (80%) and Flavobacterium psychrophilum (95%) and the microparasite Sphaerothecum destruens (53%). Fish sampled at the falls had significantly greater relative loads of Ca. B. cysticola and F. psychrophilum, higher infectious agent richness and differential gene expression compared to fish captured upstream. Flavobacterium psychrophilum was only associated with immune gene expression (particularly humoral immunity) of fish sampled at the falls, while water temperature was positively correlated with genes involved in the complement system, metabolic stress and oxidative stress for fish captured upstream. This work highlights interesting differences in agent-host interactions across fisheries and suggests that hydraulic barriers may reduce the passage of fish with the heaviest infectious agent burdens, emphasizing the selective role of areas of difficult passage. Further, this work expands our knowledge of infectious agent prevalence in wild salmonids and provides insight into the relationships between infectious agents and host physiology.

Daily Regulation of Phototransduction, Circadian Clock, DNA Repair, and Immune Gene Expression by Heme Oxygenase in the Retina of Drosophila.

The daily expression of genes and the changes in gene expression after silencing the heme oxygenase (ho) gene were examined in the retina of Drosophila using microarray and SybrGreen qPCR (quantitative polymerase chain reaction) methods. The HO decrease in the morning upregulated 83 genes and downregulated 57 genes. At night, 80 genes were upregulated and 22 were downregulated. The top 20 genes downregulated after ho silencing in the morning modulate phototransduction, immune responses, autophagy, phagocytosis, apoptosis, the carbon monoxide (CO) response, the oxidative stress/UV response, and translation. In turn, the genes that upregulated at night were involved in translation—the response to oxidative stress, DNA damage, and phototransduction. Among the top 20 genes downregulated at night were genes involved in phototransduction, immune responses, and autophagy. For some genes, a low level of HO had an opposite effect in the morning compared to those at night. Silencing ho also changed the expression of circadian clock genes, while the HO decrease during the night enhanced the expression of immune system genes. The results showed that the cyclic expression of HO is important for controlling several processes in the retina, including neuroprotection and those involved in the innate immune system.

Anti-LINGO-1 has no detectable immunomodulatory effects in preclinical and phase 1 studies.

Objective:To evaluate whether the anti-LINGO-1 antibody has immunomodulatory effects.Methods:Human peripheral blood mononuclear cells (hPBMCs), rat splenocytes, and rat CD4+ T cells were assessed to determine whether LINGO-1 was expressed and was inducible. Anti-LINGO-1 Li81 (0.1–30 μg/mL) effect on proliferation/cytokine production was assessed in purified rat CD4+ T cells and hPBMCs stimulated with antibodies to CD3 +/– CD28. In humans, the effect of 2 opicinumab (anti-LINGO-1/BIIB033; 30, 60, and 100 mg/kg) or placebo IV administrations was evaluated in RNA from blood and CSF samples taken before and after administration in phase 1 clinical trials; paired samples were assessed for differentially expressed genes by microarray. RNA from human CSF cell pellets was analyzed by quantitative real-time PCR for changes in transcripts representative of cell types, activation markers, and soluble proteins of the adaptive/innate immune systems. ELISA quantitated the levels of CXCL13 protein in human CSF supernatants.Results:LINGO-1 is not expressed in hPBMCs, rat splenocytes, or rat CD4+ T cells; LINGO-1 blockade with Li81 did not affect T-cell proliferation or cytokine production from purified rat CD4+ T cells or hPBMCs. LINGO-1 blockade with opicinumab resulted in neither significant changes in immune system gene expression in blood and CSF, nor changes in CXCL13 CSF protein levels (clinical studies).Conclusions:These data support the hypothesis that LINGO-1 blockade does not affect immune function.Classification of evidence:This study provides Class II evidence that in patients with MS, opicinumab does not have immunomodulatory effects detected by changes in immune gene transcript expression.

Abstract 4469: Targeting the tri-snRNP complex for triple-negative breast cancer therapy

Abstract Triple negative breast cancers (TNBC) account for ~15% of all breast cancers and the majority of breast cancers in women carrying a BRCA1 germline mutation. Due to the inadequacy of existing treatment, here we sought to identify shared genetic dependencies across different TNBC cell lines with the ultimate goal of uncovering novel therapeutic targets. To this end, we performed a targeted siRNA lethality screen across 11 human breast cell lines focusing on 154 genes, which we previously identified as selectively essential to a genetically-defined TNBC progenitor cell line (BPLER) relative to a myoepithelial-like isogenic cell line (HMLER). Thirty genes scored as recurrently and selectively required for survival of progenitor-like (or basal-A) TNBC lines compared to differentiated cell lines. Genes for proteins involved in RNA splicing were prominent shared dependencies among basal-A TNBC cell lines. Of particular note were those associated with the U4/U6.U5 tri-snRNP complex, including PRPF8 and PRPF38A. Transcriptome-wide RNA-seq analysis of the knockdown of either led to widespread intronic retention, expression of immune system genes and aberrant splicing of transcripts involved in protein translation, proteasome function, mitosis and apoptosis, including the TNBC dependency gene MCL1. As the proteasome and mitosis are also established TNBC vulnerabilities, we have identified specific classes of transcripts that are both particularly susceptible to perturbation of tri-snRNP activity and selectively essential for TNBC, making the tri-snRNP complex an attractive target for downstream drug development. Citation Format: Stefanie Chan, Praveen Sridhar, Ying-Jie Lock, Fabio Petrocca. Targeting the tri-snRNP complex for triple-negative breast cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4469. doi:10.1158/1538-7445.AM2017-4469