The FAM13A Long Isoform Regulates Cilia Movement and Co-ordination in Airway Mucociliary Transport.

SNPs in the FAM13A locus are amongst the most commonly reported risk alleles associated with chronic obstructive pulmonary disease (COPD) and other respiratory diseases, however the physiological role of FAM13A is unclear. In humans, two major protein isoforms are expressed at the FAM13A locus: 'long' and 'short', but their functions remain unknown, partly due to a lack of isoform conservation in mice. We performed in-depth characterisation of organotypic primary human airway epithelial cell subsets and show that multiciliated cells predominantly express the FAM13A long isoform containing a putative N-terminal Rho GTPase activating protein (RhoGAP) domain. Using purified proteins, we directly demonstrate RhoGAP activity of this domain. In Xenopus laevis, which conserve the long isoform, Fam13a-deficiency impaired cilia-dependent embryo motility. In human primary epithelial cells, long isoform deficiency did not affect multiciliogenesis but reduced cilia co-ordination in mucociliary transport assays. This is the first demonstration that FAM13A isoforms are differentially expressed within the airway epithelium, with implications for the assessment and interpretation of SNP effects on FAM13A expression levels. We also show that the long FAM13A isoform co-ordinates cilia-driven movement, suggesting that FAM13A risk alleles may affect susceptibility to respiratory diseases through deficiencies in mucociliary clearance. This article is open access and distributed under the terms of the Creative Commons Attribution 4.0 International License (https://creativecommons.org/licenses/by/4.0/).

Myeloid-derived suppressor cells in COVID-19: A review.

Coronavirus disease 2019 (COVID-19) is a potentially life-threatening infection characterized by excessive inflammation, coagulation disorders and organ damage. A dysregulated myeloid cell compartment is one of the most striking immunopathologic signatures of this newly emerged infection. A growing number of studies are reporting on the expansion of myeloid cells with immunoregulatory activities in the periphery and airways of COVID-19 patients. These cells share phenotypic and functional similarities with myeloid-derived suppressor cells (MDSCs), which were first described in cancer patients. MDSCs are a heterogeneous population of pathologically activated myeloid cells that exert immunosuppressive activities against mainly effector T cells. The increased frequency of these cells in COVID-19 patients suggests that they are involved in immune regulation during this infection. In this article, we review the current findings on MDSCs in COVID-19 and discuss the complex role of these cells in the immunopathology of COVID-19.

Fc Receptor is Involved in Nk Cell Functional Anergy Induced by Miapaca2 Tumor Cell Line.

Impaired NK cytotoxicity has been linked to poor cancer prognosis, but its mechanisms are not clearly established. Increasing data demonstrate that NK cells lose cytotoxicity after interaction with NK cell-sensitive tumor cells. In this paper, we provide evidence that the human adenocarcinoma cell line MiaPaCa2 and TNFα and TGFß-treated MiaPaCa2 cultures (MiaPaCa2-TT) induced functional anergy of NK cells via FGL2 protein. MiaPaCa2-TT cultures decreased expression of IFNγ, CD107a, DNAM-1, and stimulated expression of PD1 by NK cells, as well as inhibited their cytotoxic activity in a greater manner compared to the parental culture. More importantly, we found that co-cultivation with anergized NK cells decreased expression of IFNγ and CD107a by naïve NK cells, which supports the hypothesis of NK cell functional anergy transmission. The obtained results suggest a mechanism by which tumor cells may inhibit cytotoxic functions of tumor-infiltrating and circulating NK cells in cancer.Abbreviations: CFSE: Carboxyfluorescein diacetate succinimidyl ester; CSCs: Cancer stem cells; FGL2: Fibrinogen-like protein 2; mAbs: Monoclonal antibodies; MiaPaCa2: Human adenocarcinoma cell line; MiaPaCa2-ТТ: Adenocarcinoma cell line MiaPaCa2 cells stimulated with TNFα and TGFß-1; PI: Propidium iodide; TGFß: Transforming growth factor beta; TME: Tumor microenvironment; TNFα: Tumor necrosis factor alfa.

Rhinovirus-induced Human Lung Tissue Responses Mimic Chronic Obstructive Pulmonary Disease and Asthma Gene Signatures.

Human rhinovirus (RV) is a major risk factor for chronic obstructive pulmonary disease (COPD) and asthma exacerbations. The exploration of RV pathogenesis has been hampered by a lack of disease-relevant model systems. We performed a detailed characterization of host responses to RV infection in human lung tissue ex vivo and investigated whether these responses are disease relevant for patients with COPD and asthma. In addition, impact of the viral replication inhibitor rupintrivir was evaluated. Human precision-cut lung slices (PCLS) were infected with RV1B with or without rupintrivir. At Days 1 and 3 after infection, RV tissue localization, tissue viability, and viral load were determined. To characterize host responses to infection, mediator and whole genome analyses were performed. RV successfully replicated in PCLS airway epithelial cells and induced both antiviral and proinflammatory cytokines such as IFNα2a, CXCL10, CXCL11, IFN-γ, TNFα, and CCL5. Genomic analyses revealed that RV not only induced antiviral immune responses but also triggered changes in epithelial cell-associated pathways. Strikingly, the RV response in PCLS was reflective of gene expression changes described in patients with COPD and asthma. Although RV-induced host immune responses were abrogated by rupintrivir, RV-triggered epithelial processes were largely refractory to antiviral treatment. Detailed analysis of RV-infected human PCLS and comparison with gene signatures of patients with COPD and asthma revealed that the human RV PCLS model represents disease-relevant biological mechanisms that can be partially inhibited by a well-known antiviral compound and provide an outstanding opportunity to evaluate novel therapeutics.

Oral administration of ammonium metavanadate and potassium dichromate distorts the inflammatory reaction induced by turpentine oil injection in male rats.

Heavy metal pollution is rapidly increasing in the environment. It has been shown that exposure to vanadium and chromium is able to alter the immune response. Nevertheless, the mechanisms by which these metal pollutants mediate their immunomodulatory effects are not completely understood. Herein, we examined the effect of ammonium metavanadate and potassium dichromate on the development of an inflammatory response caused by subcutaneous injection of turpentine oil. We demonstrated that pretreatment of rats with ammonium metavanadate and potassium dichromate for two weeks prior to initiation of the inflammatory response resulted in a wider zone of necrosis surrounding the site of inflammation. The acute inflammatory process in the combined model was characterized by elevated serum levels of IL-10 and decreased serum levels of IL-6 as compared to rats not treated with ammonium metavanadate and potassium dichromate. Ammonium metavanadate and potassium dichromate administration induced a decrease in the proportion of splenic His48HighCD11b/c+ myeloid cells accompanied by a reduced infiltration of the wound with neutrophils. Further analysis showed decreased proportions of CD3+CD4+IFNγ+ and CD3+CD4+IL-4+ T cells in the rats with combined model as compared to inflamed rats not treated with ammonium metavanadate and potassium dichromate. The data suggest that consumption of vanadium and chromium compounds disrupts the inflammatory response through an altered balance of pro- and anti-inflammatory cytokines and inhibition of effector T cell activation and neutrophil expansion.

Microbial Processes and Microbial Communities in the Water Column of the Polar Meromictic Lake Bol'shie Khruslomeny at the White Sea Coast.

Microbiological, molecular ecological, biogeochemical, and isotope geochemical research was carried out at the polar Lake Bol'shie Khruslomeny at the coast of the Kandalaksha Bay, White Sea in March and September 2017. The uppermost mixolimnion was oxic, with low salinity (3-5%). The lower chemocline layer was brown-green colored, with very high content of particulate organic matter (up to 11.8 mg C L-1). The lowermost monimolimnion had marine salinity (22-24%) and very high concentrations of sulfide (up to 18 mmol L-1) and CH4 (up to 1.8 mmol L-1). In the chemocline, total microbial abundance and the rate of anoxygenic photosynthesis were 8.8 × 106 cells mL-1 and 34.4 µmol C L-1 day-1, respectively. Both in March and September, sulfate reduction rate increased with depth, peaking (up to 0.6-1.1 µmol S L-1 day-1) in the lower chemocline. Methane oxidation rates in the chemocline were up to 85 and 180 nmol CH4 L-1 day-1 in March and September, respectively; stimulation of this process by light was observed in September. The percentages of cyanobacteria and methanotrophs in the layer where light-induced methane oxidation occurred were similar, ∼2.5% of the microbial community. Light did not stimulate methane oxidation in deeper layers. The carbon isotope composition of particulate organic matter (δ13C-Corg), dissolved carbonates (δ13C-DIC), and methane (δ13C- CH4) indicated high microbial activity in the chemocline. Analysis of the 16S rRNA gene sequences revealed predominance of Cyanobium cyanobacteria (order Synechococcales) in the mixolimnion. Green sulfur bacteria Chlorobium phaeovibrioides capable of anoxygenic photosynthesis constituted ∼20% of the chemocline community both in March and in September. Methyloprofundus gammaptoteobacteria (family Methylomonaceae) were present in the upper chemocline, where active methane oxidation occurred. During winter, cyanobacteria were less abundant in the chemocline, while methanotrophs occurred in higher horizons, including the under-ice layer. Chemolithotrophic gammaproteobacteria of the genus Thiomicrorhabdus, oxidizing reduced sulfur compounds at low oxygen concentrations, were revealed in the chemocline in March. Both in March and September archaea constituted up to 50% of all microorganisms in the hypolimnion. The percentage of putative methanogens in the archaeal community was low, and they occurred mainly in near-bottom horizons.

Exogenous Melatonin Up-Regulates Expression of CD62L by Lymphocytes in Aged Mice under Inflammatory and Non-Inflammatory Conditions.

It is well documented that age-related impaired functioning of immunocompetent cells is associated with an increase in the rates of chronic inflammatory diseases. Recently, an ability of melatonin to modulate inflammatory processes by regulating leucocyte recruitment has been demonstrated. However, to date, no studies have attempted to determine the impact of melatonin on the expression of CD62L by lymphocytes. CD62L, also known as L-selectin, is required for the entry of lymphocytes into secondary lymphoid organs, sites of tumor growth and chronic inflammation through high endothelial venules. Here, we investigated the effect of melatonin at physiological concentrations on the expression of CD62L by T and NK cells in vivo and in vitro. We demonstrated that NK and CD3+ T cells obtained from the spleen of aged mice were characterized by decreased expression of CD62L compared to young mice. Melatonin administration up-regulated the levels of surface CD62L on NK and T cell populations in aged mice under non-inflammatory conditions and on CD8+ T cells in aged mice with chronic inflammation. Pre-incubation with melatonin prevented the reduction in CD62L expression by CD8+ T cells induced by the co-cultivation of peripheral blood mononuclear cells with human pancreatic adenocarcinoma cell line (MiaPaCa-2). The obtained results suggest that melatonin can modulate lymphocyte homing into lymph nodes and sites of chronic inflammation and, therefore, can stimulate immune responses in chronic inflammatory conditions associated with aging.

Chronic Inflammation Contributes to Tumor Growth: Possible Role of L-Selectin-Expressing Myeloid-Derived Suppressor Cells (MDSCs).

Recent data have demonstrated that chronic inflammation is a crucial component of tumor initiation and progression. We previously reported that immature myeloid-derived suppressor cells (MDSCs) with immunosuppressive activity toward effector T cells were expanded in experimental chronic inflammation. We hypothesized that elevated levels of MDSCs, induced by chronic inflammation, may contribute to the progression of tumor growth. Using the Ehrlich carcinoma animal model, we found increased tumor growth in mice with chronic adjuvant arthritis, which was accompanied by a persistent increase in the proportion of splenic monocytic and granulocytic MDSCs expressing CD62L (L-selectin), when compared to tumor mice without adjuvant arthritis. Depletion of inflammation-induced MDSCs resulted in decreased tumor growth. In vitro studies demonstrated that increased expression of CD62L by MDSCs was mediated by TNFα, elevated concentrations of which were found in tumor mice subjected to chronic inflammation. Moreover, the addition of exogenous TNFα markedly enhanced the suppressive activity of bone marrow-derived MDSCs, as revealed by the ability to impair the proliferation of CD8+ T cells in vitro. This study provides evidence that chronic inflammation may promote tumor growth via induction of CD62L expression by MDSCs that can facilitate their migration to tumor and lymph nodes and modulation of their suppressor activity.

Modulation of the expression of genes related to the system of amyloid-beta metabolism in the brain as a novel mechanism of ceftriaxone neuroprotective properties.

BACKGROUND: The dominant hypothesis about the pathogenesis of Alzheimer's disease (AD) is the "amyloid cascade" concept and modulating the expression of proteins involved in the metabolism of amyloid-beta (Aß) is proposed as an effective strategy for the prevention and therapy of AD. Recently, we found that an antibiotic ceftriaxone (CEF), which possesses neuroprotective activity, reduced cognitive deficits and neurodegenerative changes in OXYS rats, a model of sporadic AD. The molecular mechanisms of this effect are not completely clear, we suggested that the drug might serve as the regulator of the expression of the genes involved in the metabolism of Aß and the pathogenesis of AD. The study was aimed to determine the effects of CEF on mRNA levels of Bace1 (encoding ß-secretase BACE1 involved in Aß production), Mme, Ide, Ece1, Ace2 (encoding enzymes involved in Aß degradation), Epo (encoding erythropoietin related to endothelial function and clearance of Aß across the blood brain barrier) in the frontal cortex, hippocampus, striatum, hypothalamus, and amygdala of OXYS and Wistar (control strain) male rats. Starting from the age of 14 weeks, animals received CEF (100 mg/kg/day, i.p., 36 days) or saline. mRNA levels were evaluated with RT-qPCR method. Biochemical parameters of plasma were measured for control of system effects of the treatment. RESULTS: To better understand strain variations studied here, we compared the gene expression between untreated OXYS and Wistar rats. This comparison showed a significant decrease in mRNA levels of Ace2 in the frontal cortex and hypothalamus, and of Actb in the amygdala of untreated OXYS rats. Analysis of potential effects of CEF revealed its novel targets. In the compound-treated OXYS cohort, CEF diminished mRNA levels of Bace1 and Ace2 in the hypothalamus, and Aktb in the frontal cortex. Furthermore, CEF augmented Mme, Ide, and Epo mRNA levels in the amygdala as well as the levels of Ece1 and Aktb in the striatum. Finally, CEF also attenuated the activity of ALT and AST in plasma of OXYS rats. CONCLUSION: Those findings disclosed novel targets for CEF action that might be involved into neuroprotective mechanisms at early, pre-plaque stages of AD-like pathology development.

Functional heterogeneity of circulating T regulatory cell subsets in breast cancer patients.

BACKGROUND: Regulatory T cells (Tregs) play a major role in tumor escape from immunosurveillance by suppressing effector cells. The number of Tregs is increased in tumor sites and peripheral blood of breast cancer patients. However, the data regarding phenotypic and functional heterogeneity of Treg subpopulations in breast cancer are limited. The present study aimed to investigate the number and suppressive potential of Tregs that possess natural naïve-(N nTregs), effector/memory-like (EM nTregs), and Tr1-like phenotypes in breast cancer patients and healthy women. METHODS: The study included 10 HW and 17 primary breast cancer patients. Numbers of CD4+CD25+FoxP3+CD45RA+ N nTregs, CD4+CD25+FoxP3+CD45RA- EM nTregs, and CD4+IL-4-IL-10+ Tr1 subsets and the expression of CTLA-4, CD39, GITR, LAP, and IL-35 by these Treg subsets were measured in freshly obtained peripheral blood by flow cytometry. RESULTS: Herein, we demonstrate that the percentages of N nTregs, EM nTregs, CD25+ and FoxP3+ Tr1 cells are elevated in the peripheral blood of breast cancer patients, but do not correlate with cancer stages. Nevertheless, the frequency of CD25+ Tr1 cells was associated with nodal involvement, while the number of EM nTregs correlated with clinical outcome. The expression of CTLA-4 and IL-35 by all assessed Treg subsets was increased throughout all tumor stages (I-III). CONCLUSIONS: Collectively, the current study shows phenotypic alterations in suppressive receptors of Treg subsets, suggesting that breast cancer patients have increased activity of N nTregs, EM nTregs and Tr1 cells; and EM nTregs and CD25+ Tr1 cells represent prospective markers for assessing disease prognosis.

Daunorubicin conjugated with alpha-fetoprotein selectively eliminates myeloid-derived suppressor cells (MDSCs) and inhibits experimental tumor growth.

Failure of antitumor immunity in cancer was shown to be mediated by myeloid-derived suppressor cells (MDSCs), which are considered to be one of the key factors contributing to the development of malignant diseases. Therefore, the development of pharmacological approaches to effectively eliminate MDSCs in organisms carrying growing tumors is a promising pathway for potential treatment. For this purpose we propose alpha-fetoprotein (AFP) conjugated with a cytotoxic agent as a vector molecule, specifically recognizing MDSCs. The present study was aimed at examination of this suggestion using both in vitro and in vivo approaches. MDSCs, obtained from the spleen of Ehrlich carcinoma bearing mice, selectively bound AFP labeled with fluorescein isothiocyanate. AFP conjugated to daunorubicin (AFP-DR) and DR alone showed similar in vitro cytotoxicity against the granulocytic MDSC subpopulation. The monocytic MDSC subpopulation was resistant to treatment with DR, whereas it was completely depleted in the presence of AFP-DR. Treatment of mice bearing Ehrlich carcinoma with AFP-DR resulted in reduced numbers of splenic MDSCs, normalization of NK cell levels, and inhibition of tumor growth. The obtained results demonstrate that cytotoxic conjugates based on AFP are promising anticancer drugs, which, in addition to the direct effect on tumor cells expressing receptors to AFP, may contribute to elimination of MDSCs.

TNFα and TGFß-1 synergistically increase the cancer stem cell properties of MiaPaCa-2 cells.

Increased serum concentrations of tumor necrosis factor α (TNFα) and transforming growth factor ß-1 (TGFß-1) in the blood of patients with pancreatic cancer (PC) have previously been demonstrated. In addition, exogenous exposure to these cytokines promotes various cancer cell invasive and cancer stem cell (CSC) phenotypes. However, their importance in pancreatic CSCs remains elusive. In the present study, the effects of TNFα and TGFß-1 on the human PC cell line MiaPaCa-2 were examined. Using flow cytometry, it was revealed that TNFα and TGFß-1 synergistically increase cluster of differentiation (CD) 44v6, CD133 and ATP-binding cassette transporter G2 (ABCG2) expressing populations in adherent tumor cell culture conditions. Furthermore, a similar trend was observed in cells pretreated with these cytokines grown in sphere forming culture conditions. Similar to previous studies, TNFα treatment increased the proportion of epidermal growth factor receptor (EGFR) expressing cells in adherent culture, and this data was further supported by the results of the sphere formation assay, in which the subculture with a high proportion of EGFR expressing cells exhibited the most efficient sphere forming ability. However, the proportion of vascular endothelial growth factor receptor 1 (VEGFR1) expressing cells did not increase upon treatment with these cytokines individually or in combination. This data was subsequently supported by the results of the wound healing assay in which cytokine treatment did not increase the migration of cells. The MTT cell proliferation and cytotoxicity assay revealed that TNFα + TGFß-1 treatment significantly increased cell proliferation and daunorubicin resistance, but not gemcitabine resistance. In conclusion, the data of the current study provide a mechanistic association between TNFα, TGFß-1 and the CSC properties of MiaPaCa-2 cells. In addition, it suggests that targeting TNFα and TGFß-1 is beneficial for improving the therapeutic efficacy of treatments for patients with PC.

Expansion of CD11b+Ly6Ghigh and CD11b+CD49d+ myeloid cells with suppressive potential in mice with chronic inflammation and light-at-night-induced circadian disruption.

OBJECTIVE: Myeloid-derived suppressor cells (MDSCs) are important negative regulators of immune processes in cancer and other pathological conditions. We suggested that MDSCs play a key role in pathogenesis of chronic inflammation, which precedes and, to a certain extent, induces carcinogenesis. The present study aimed at investigation of MDSCs arising during chronic inflammation and light-at-night (LN)-induced stress, which is shown to accelerate chronic diseases. SUBJECTS: 67 CD-1 mice and in vitro MDSC cultures. TREATMENT: Adjuvant arthritis was induced by a subdermal injection of complete Freund's adjuvant. LN was induced by illumination of 750 lx at night. METHODS: Flow cytometry for evaluation of cell phenotypes and MTT standard test for cell proliferation were used. RESULTS: Increased levels of splenic CD11b+Ly6Ghigh and CD11b+CD49d+ myeloid cells possessing suppressive potential in mice with adjuvant arthritis are shown. LN amplifies the process of CD11b+Ly6Ghigh expansion in mice with adjuvant arthritis. Expression of CD62L and CD195 is elevated on the myeloid cells during exposure to LN. CONCLUSIONS: Our study raises the possibility that CD11b+Ly6Ghigh and CD11b+CD49d+ MDSCs play an important role in the induction of immunosuppressive environment typical for chronic inflammation. Also, LN can affect immune responses during chronic inflammation through recruitment of MDSCs from the bone marrow.

New Insights into Epigenetic and Pharmacological Regulation of Amyloid-Degrading Enzymes.

Currently, deficit of amyloid ß-peptide (Aß) clearance from the brain is considered as one of the possible causes of amyloid accumulation and neuronal death in the sporadic form of Alzheimer's disease (AD). Aß clearance can involve either specific proteases present in the brain or Aß-binding/transport proteins. Among amyloid-degrading enzymes the most intensively studied are neprilysin (NEP) and insulin-degrading enzyme (IDE). Since ageing and development of brain pathologies is often accompanied by a deficit in the levels of expression and activity of these enzymes in the brain, there is an urgent need to understand the mechanisms involved in their regulation. We have recently reported that NEP and also an Aß-transport protein, transthyretin are epigenetically co-regulated by the APP intracellular domain (AICD) and this regulation depends on the cell type and APP695 isoform expression in a process that can be regulated by the tyrosine kinase inhibitor, Gleevec. We have now extended our work and shown that, unlike NEP, another amyloid-degrading enzyme, IDE, is not related to over-expression of APP695 in neuroblastoma SH-SY5Y cells but is up-regulated by APP751 and APP770 isoforms independently of AICD but correlating with reduced HDAC1 binding to its promoter. Studying the effect of the nuclear retinoid X receptor agonist, bexarotene, on NEP and IDE expression, we have found that both enzymes can be up-regulated by this compound but this mechanism is not APP-isoform specific and does not involve AICD but, on the contrary, affects HDAC1 occupancy on the NEP gene promoter. These new insights into the mechanisms of NEP and IDE regulation suggest possible pharmacological targets in developing AD therapies.

Peripheral blood NK cells expressing HLA-G, IL-10 and TGF-ß in healthy donors and breast cancer patients.

Human natural killer (NK) cells are not only professional cytotoxic cells integrated into effector branch of innate immunity, but they are also regulatory cells, managing different immune processes. Immunoregulatory NK cells, expressing HLA-G and IL-10, have been generated in vitro from human hematopoietic progenitors and found in vivo among decidual NK cells of pregnant women. Human peripheral blood NK cells have been shown to acquire suppressive properties after HLA-G uptake during trogocytosis. Moreover, it has been shown that circulating NK cells contain a trace amount of cells producing TGF-ß and IL-10, which exert a suppressive influence upon innate and adaptive immunity. In this study, we report on a minor subset of peripheral blood HLA-G(+) NK cells possessing suppressive activity toward effector functions of NK cells. Further we demonstrate an increased number of circulating HLA-G(+), IL-10(+), and TGF-ß(+) NK cells in breast cancer patients which might impair efficiency of anti-tumor immunity.

Amyloid-clearing proteins and their epigenetic regulation as a therapeutic target in Alzheimer's disease.

Abnormal elevation of amyloid ß-peptide (Aß) levels in the brain is the primary trigger for neuronal cell death specific to Alzheimer's disease (AD). It is now evident that Aß levels in the brain are manipulable due to a dynamic equilibrium between its production from the amyloid precursor protein (APP) and removal by amyloid clearance proteins. Clearance can be either enzymic or non-enzymic (binding/transport proteins). Intriguingly several of the main amyloid-degrading enzymes (ADEs) are members of the M13 peptidase family (neprilysin (NEP), NEP2 and the endothelin converting enzymes (ECE-1 and -2)). A distinct metallopeptidase, insulin-degrading enzyme (IDE), also contributes to Aß degradation in the brain. The ADE family currently embraces more than 20 members, both membrane-bound and soluble, and of differing cellular locations. NEP plays an important role in brain function terminating neuropeptide signals. Its decrease in specific brain areas with age or after hypoxia, ischaemia or stroke contribute significantly to the development of AD pathology. The recently discovered mechanism of epigenetic regulation of NEP (and other genes) by the APP intracellular domain (AICD) and its dependence on the cell type and APP isoform expression suggest possibilities for selective manipulation of NEP gene expression in neuronal cells. We have also observed that another amyloid-clearing protein, namely transthyretin (TTR), is also regulated in the neuronal cell by a mechanism similar to NEP. Dependence of amyloid clearance proteins on histone deacetylases and the ability of HDAC inhibitors to up-regulate their expression in the brain opens new avenues for developing preventive strategies in AD.

The Aß-clearance protein transthyretin, like neprilysin, is epigenetically regulated by the amyloid precursor protein intracellular domain.

Proteolytic cleavage of the amyloid precursor protein (APP) by the successive actions of ß- and γ-secretases generates several biologically active metabolites including the amyloid ß-peptide (Aß) and the APP intracellular domain (AICD). By analogy with the Notch signalling pathway, AICD has been proposed to play a role in transcriptional regulation. Among the cohort of genes regulated by AICD is the Aß-degrading enzyme neprilysin (NEP). AICD binds to the NEP promoter causing transcriptional activation by competitive replacement with histone deacetylases (HDACs) leading to increased levels of NEP activity and hence increased Aß clearance. We now show that the Aß-clearance protein transthyretin (TTR) is also epigenetically up-regulated by AICD. Like NEP regulation, AICD derived specifically from the neuronal APP isoform, APP695 , binds directly to the TTR promoter displacing HDAC1 and HDAC3. Cell treatment with the tyrosine kinase inhibitor Gleevec (imatinib) or with the alkalizing agent NH4 Cl causes an accumulation of 'functional' AICD capable of up-regulating both TTR and NEP, leading to a reduction in total cellular Aß levels. Pharmacological regulation of both NEP and TTR might represent a viable therapeutic target in Alzheimer's disease.

Transmission of "split anergy" from tumor infiltrating to peripheral NK cells in a manner similar to "infectious tolerance".

According to a new paradigm of carcinogenesis, a tumor arises not from transformed cell, but only from tumor initiating cells called cancer stem cells (CSCs), which can originate from tissue stem cells. CSC are resistant to conventional therapy and after treatment form new tumors and give rise to metastases. Only natural killer (NK) cells are capable of lysing CSCs, but within different tumor types these cells experience a condition known as "split anergy", whereby the NK cells lose the ability to kill CSCs and being to produce cytokines. As a result, uncontrolled tumor growth arises and tumor stroma accumulates anergic NK cells. We hypothesize that anergic tumor infiltrating NK (TINK) cells transmit their property to naïve NK cells by infecting" them with a state of "split anergy" in a similar manner as T conventional cells are transformed into T regulatory cells during the process of "infectious tolerance". Anergic TINK cells egress from the tumor stroma via the lymphatic system, where they reach regional lymph nodes and transmit their properties to naïve NK cells, which in turn become anergic toward CSCs and lose immunosurveillance functions. The mechanisms proposed for this hypothesis and the methodological approaches for confirming the idea are presented in this issue.

Epigenetic regulation of angiotensin-converting enzyme 2 (ACE2) by SIRT1 under conditions of cell energy stress.

ACE2 (angiotensin-converting enzyme 2) counterbalances the actions of ACE (angiotensin-converting enzyme) by metabolizing its catalytic product, the vasoactive and fibrogenic peptide AngII (angiotensin II), into Ang-(1-7) [angiotensin-(1-7)]. Enhanced ACE2 expression may be protective in diabetes, cardiovascular disease and cancer. However, relatively little is known about the specific physiological factors regulating ACE2 expression. In the present paper, we show, by Western blotting and qPCR (quantitative real-time PCR), that ACE2 expression is increased under conditions of cell stress, including hypoxic conditions, IL (interleukin)-1ß treatment and treatment with the AMP mimic AICAR (5-amino-4-imidazolecarboxamide riboside). The NAD+-dependent deacetylase SIRT1 (silent information regulator T1) was found to be up-regulated after AICAR treatment but, conversely, was down-regulated after IL-1ß treatment. ChIP analysis demonstrated that SIRT1 bound to the ACE2 promoter and that binding was increased after AICAR treatment, but decreased after IL-1ß treatment. Inhibition of SIRT1 activity ablated the AICAR-induced increase in ACE2. In conclusion, we have established that the expression of the ACE2 transcript is controlled by the activity of SIRT1 under conditions of energy stress.