Akt Phosphorylation Influences Persistent Chlamydial Infection and Chlamydia-Induced Golgi Fragmentation Without Involving Rab14

Chlamydia trachomatis is an obligate intracellular bacterium that causes multiple diseases involving the eyes, gastrointestinal tract, and genitourinary system. Previous studies have identified that in acute chlamydial infection, C. trachomatis requires Akt pathway phosphorylation and Rab14-positive vesicles to transmit essential lipids from the Golgi apparatus in survival and replication. However, the roles that Akt phosphorylation and Rab14 play in persistent chlamydial infection remain unclear. Here, we discovered that the level of Akt phosphorylation was lower in persistent chlamydial infection, and positively correlated with the effect of activating the development of Chlamydia but did not change the infectivity and 16s rRNA gene expression. Rab14 was found to exert a limited effect on persistent infection. Akt phosphorylation might regulate Chlamydia development and Chlamydia-induced Golgi fragmentation in persistent infection without involving Rab14. Our results provide a new insight regarding the potential of synergistic repressive effects of an Akt inhibitor with antibiotics in the treatment of persistent chlamydial infection induced by penicillin.

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Use of Drosophila S2 Cells as a Model for Studying Ehrlichia chaffeensis Infections

Applied and Environmental Microbiology ◽

10.1128/aem.02467-07 ◽

2008 ◽

Vol 74 (6) ◽

pp. 1886-1891 ◽

Cited By ~ 14

Author(s):

Alison Luce-Fedrow ◽

Tonia Von Ohlen ◽

Daniel Boyle ◽

Roman R. Ganta ◽

Stephen K. Chapes

Keyword(s):

Mammalian Cells ◽

Genetic Manipulation ◽

Rrna Gene ◽

Ehrlichia Chaffeensis ◽

S2 Cells ◽

Obligate Intracellular Bacterium ◽

Obligate Intracellular ◽

Mammalian Cell Lines ◽

Drosophila S2 Cells ◽

Human Monocytic Ehrlichiosis

ABSTRACT Ehrlichia chaffeensis is an obligate intracellular bacterium and the causative agent of human monocytic ehrlichiosis. Although this pathogen grows in several mammalian cell lines, no general model for eukaryotic cellular requirements for bacteria replication has yet been proposed. We found that Drosophila S2 cells are permissive for the growth of E. chaffeensis. We saw morulae (aggregates of bacteria) by microscopy, detected the E. chaffeensis 16S rRNA gene by reverse transcriptase PCR, and used immunocytochemistry to detect E. chaffeensis in S2 and mammalian cells. Bacteria grown in S2 cells reinfected mammalian macrophages. S2 cells were made nonpermissive for E. chaffeensis through incubation with lipopolysaccharide. Our results demonstrate that S2 cells are an appropriate system for studying the pathogenesis of E. chaffeensis. The use of a Drosophila system has the potential to serve as a model system for studying Ehrlichia due to its completed genome, ease of genetic manipulation, and the availability of mutants.

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Suppression of Sirtuin-1 Increases IL-6 Expression by Activation of the Akt Pathway During Allergic Asthma

Cellular Physiology and Biochemistry ◽

10.1159/000484119 ◽

2017 ◽

Vol 43 (5) ◽

pp. 1950-1960 ◽

Cited By ~ 19

Author(s):

Lingling Tang ◽

Qingge Chen ◽

Ziyu Meng ◽

Li Sun ◽

Linyun Zhu ◽

...

Keyword(s):

Allergic Asthma ◽

Sirtuin 1 ◽

Akt Pathway ◽

Epithelial Cell Line ◽

Dependent Manner ◽

Airway Epithelial ◽

Mice Model ◽

Akt Inhibitor ◽

Akt Phosphorylation ◽

Protein Levels

Background/Aims: A growing number of studies have demonstrated that the activity and expression level of sirtuin-1 (SIRT1) are decreased in asthma patients; however, the mechanisms underlying decreased SIRT1 expression and function are still not completely understood. Interleukin (IL)-6 plays important roles in inflammation during allergic asthma. In this study, we examined whether loss of SIRT1 activity regulated the expression of IL-6 and further verified the underlying mechanisms. Methods: The human airway epithelial cell line 16HBE was used to test the effects of the SIRT1 inhibitor (salermide) on expression of IL-6. IL-6 mRNA and protein expression were assessed with real-time polymerase chain reaction (PCR), immunochemistry, and ELISA. OVA-challenged mice were used as an asthma model to investigate the effect of SIRT1 activation on IL-6 and relative Akt phosphorylation level. Results: We found that inhibition of SIRT1 increased IL-6 mRNA and protein levels in a time-dependent manner, which was accompanied by increased Akt pathway activation in 16HBE cells. Furthermore activation of Akt showed upregulated expression of the IL-6 protein whereas Akt inhibitor, LY294002 or Akt siRNA significantly inhibited SIRT1-regulated IL-6 expression. Conversely, activation of SIRT1 inhibited Akt activation and IL-6 expression in an asthmatic mice model and 16HBE cells. Conclusion: Our results indicate the potential role of SIRT1 in regulating inflammation by modulation of IL-6 expression in an Akt-dependent manner during allergic asthma.

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Physical and Genetic Map of the Obligate Intracellular Bacterium Coxiella burnetii

Journal of Bacteriology ◽

10.1128/jb.180.15.3816-3822.1998 ◽

1998 ◽

Vol 180 (15) ◽

pp. 3816-3822 ◽

Cited By ~ 28

Author(s):

H. Willems ◽

Cornelie Jäger ◽

Georg Baljer

Keyword(s):

Genetic Map ◽

Coxiella Burnetii ◽

Physical Map ◽

Gene Organization ◽

Intracellular Bacterium ◽

Rrna Gene ◽

Obligate Intracellular Bacterium ◽

Obligate Intracellular ◽

Great Heterogeneity ◽

Physical And Genetic Map

ABSTRACT Pulsed-field gel electrophoresis and PCR techniques have been used to construct a NotI macrorestriction map of the obligate intracellular bacterium Coxiella burnetii Nine Mile. The size of the chromosome has been determined to be 2,103 kb comprising 29NotI restriction fragments. The average resolution is 72.5 kb, or about 3.5% of the genome. Experimental data support the presence of a linear chromosome. Published genes were localized on the physical map by Southern hybridization. One gene, recognized as transposable element, was found to be present in at least nine sites evenly distributed over the whole chromosome. There is only one copy of a 16S rRNA gene. The putative oriC has been located on a 27.5-kb NotI fragment. Gene organization upstream theoriC is almost identical to that of Pseudomonas putida and Bacillus subtilis, whereas gene organization downstream the oriC seems to be unique among bacteria. The physical map will be helpful in investigations of the great heterogeneity in restriction fragment length polymorphism patterns of different isolates and the great variation in genome size. The genetic map will help to determine whether gene order in different isolates is conserved.

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Combined Application of Pan-AKT Inhibitor MK-2206 and BCL-2 Antagonist Venetoclax in B-Cell Precursor Acute Lymphoblastic Leukemia

International Journal of Molecular Sciences ◽

10.3390/ijms22052771 ◽

2021 ◽

Vol 22 (5) ◽

pp. 2771

Author(s):

Anna Richter ◽

Elisabeth Fischer ◽

Clemens Holz ◽

Julia Schulze ◽

Sandra Lange ◽

...

Keyword(s):

Cell Proliferation ◽

Tumor Cell ◽

Cell Lines ◽

Morphological Changes ◽

Synergistic Effects ◽

Lymphoblastic Leukemia ◽

Tumor Cell Proliferation ◽

Akt Inhibitor ◽

Akt Phosphorylation ◽

Combined Application

Aberrant PI3K/AKT signaling is a hallmark of acute B-lymphoblastic leukemia (B-ALL) resulting in increased tumor cell proliferation and apoptosis deficiency. While previous AKT inhibitors struggled with selectivity, MK-2206 promises meticulous pan-AKT targeting with proven anti-tumor activity. We herein, characterize the effect of MK-2206 on B-ALL cell lines and primary samples and investigate potential synergistic effects with BCL-2 inhibitor venetoclax to overcome limitations in apoptosis induction. MK-2206 incubation reduced AKT phosphorylation and influenced downstream signaling activity. Interestingly, after MK-2206 mono application tumor cell proliferation and metabolic activity were diminished significantly independently of basal AKT phosphorylation. Morphological changes but no induction of apoptosis was detected in the observed cell lines. In contrast, primary samples cultivated in a protective microenvironment showed a decrease in vital cells. Combined MK-2206 and venetoclax incubation resulted in partially synergistic anti-proliferative effects independently of application sequence in SEM and RS4;11 cell lines. Venetoclax-mediated apoptosis was not intensified by addition of MK-2206. Functional assessment of BCL-2 inhibition via Bax translocation assay revealed slightly increased pro-apoptotic signaling after combined MK-2206 and venetoclax incubation. In summary, we demonstrate that the pan-AKT inhibitor MK-2206 potently blocks B-ALL cell proliferation and for the first time characterize the synergistic effect of combined MK-2206 and venetoclax treatment in B-ALL.

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Assessment of PI3K/mTOR/AKT Pathway Elements to Serve as Biomarkers and Therapeutic Targets in Penile Cancer

Cancers ◽

10.3390/cancers13102323 ◽

2021 ◽

Vol 13 (10) ◽

pp. 2323

Author(s):

Anita Thomas ◽

Sascha Reetz ◽

Philipp Stenzel ◽

Katrin Tagscherer ◽

Wilfried Roth ◽

...

Keyword(s):

Cell Viability ◽

Cell Lines ◽

Penile Cancer ◽

Multivariate Regression Analysis ◽

Clinicopathological Characteristics ◽

Akt Pathway ◽

Akt Inhibitor ◽

Free Survival ◽

Kaplan Meier ◽

Treatment Naïve

The PI3K/mTOR/AKT pathway might represent an intriguing option for treatment of penile cancer (PeCa). We aimed to assess whether members of this pathway might serve as biomarkers and targets for systemic therapy. Tissue of primary cancer from treatment-naïve PeCa patients was used for tissue microarray analysis. Immunohistochemical staining was performed with antibodies against AKT, pAKT, mTOR, pmTOR, pS6, pPRAS, p4EBP1, S6K1 and pp70S6K. Protein expression was correlated with clinicopathological characteristics as well as overall survival (OS), disease-specific survival (DSS), recurrence-free survival (RFS) and metastasis-free survival (MFS). AKT inhibition was tested in two primarily established, treatment-naïve PeCa cell lines by treatment with capivasertib and analysis of cell viability and chemotaxis. A total of 76 patients surgically treated for invasive PeCa were included. Higher expression of AKT was significantly more prevalent in high-grade tumors and predictive of DSS and OS in the Kaplan–Meier analysis, and an independent predictor of worse OS and DSS in the multivariate regression analysis. Treatment with pan-AKT inhibitor capivasertib in PeCa cell lines induced a significant downregulation of both total AKT and pAKT as well as decreased cell viability and chemotaxis. Selected protein candidates of the mTOR/AKT signaling pathway demonstrate association with histological and survival parameters of PeCa patients, whereas AKT appears to be the most promising one.

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CXCR2 Blockade Influences Anaplasma phagocytophilum Propagation but Not Histopathology in the Mouse Model of Human Granulocytic Anaplasmosis

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.11.5.963-968.2004 ◽

2004 ◽

Vol 11 (5) ◽

pp. 963-968 ◽

Cited By ~ 34

Author(s):

Diana G. Scorpio ◽

Mustafa Akkoyunlu ◽

Erol Fikrig ◽

J. Stephen Dumler

Keyword(s):

Anaplasma Phagocytophilum ◽

Tissue Injury ◽

Treated Group ◽

Obligate Intracellular Bacterium ◽

Human Granulocytic Anaplasmosis ◽

Obligate Intracellular ◽

Liver Histopathology ◽

Granulocytic Anaplasmosis ◽

Infected Cells ◽

And Control

ABSTRACT Anaplasma phagocytophilum is an obligate intracellular bacterium that infects neutrophils and causes human granulocytic anaplasmosis. Infection induces neutrophil secretion of interleukin-8 or murine homologs and perpetuates infection by recruiting susceptible neutrophils. We hypothesized that antibody blockade of CXCR2 would decrease A. phagocytophilum tissue load by interrupting neutrophil recruitment but would not influence murine hepatic pathology. C3H-scid mice were treated with CXCR2 antiserum or control prior to or on day 14 after infection. Quantitative PCR and immunohistochemistry for A. phagocytophilum were performed and severity of liver histopathology was ranked. Control mice had more infected cells in tissues than the anti-CXCR2-treated group. The histopathological rank was not different between treated and control animals. Infected cells of control mice clustered in tissue more than in treated mice. The results support the hypothesis of bacterial propagation through chemokine induction and confirm that tissue injury is unrelated to A. phagocytophilum tissue load.

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Monophosphoryl lipid A induces protection against LPS in medullary thick ascending limb through a TLR4-TRIF-PI3K signaling pathway

AJP Renal Physiology ◽

10.1152/ajprenal.00064.2017 ◽

2017 ◽

Vol 313 (1) ◽

pp. F103-F115 ◽

Cited By ~ 10

Author(s):

Bruns A. Watts ◽

Thampi George ◽

Edward R. Sherwood ◽

David W. Good

Keyword(s):

Bacterial Infections ◽

Lipid A ◽

Thick Ascending Limb ◽

Akt Inhibitor ◽

Erk Activation ◽

Akt Phosphorylation ◽

Medullary Thick Ascending Limb ◽

Monophosphoryl Lipid A ◽

Monophosphoryl Lipid

Monophosphoryl lipid A (MPLA) is a detoxified derivative of LPS that induces tolerance to LPS and augments host resistance to bacterial infections. Previously, we demonstrated that LPS inhibits [Formula: see text] absorption in the medullary thick ascending limb (MTAL) through a basolateral Toll-like receptor 4 (TLR4)-myeloid differentiation factor 88 (MyD88)-ERK pathway. Here we examined whether pretreatment with MPLA would attenuate LPS inhibition. MTALs from rats were perfused in vitro with MPLA (1 µg/ml) in bath and lumen or bath alone for 2 h, and then LPS was added to (and MPLA removed from) the bath solution. Pretreatment with MPLA eliminated LPS-induced inhibition of [Formula: see text] absorption. In MTALs pretreated with MPLA plus a phosphatidylinositol 3-kinase (PI3K) or Akt inhibitor, LPS decreased [Formula: see text] absorption. MPLA increased Akt phosphorylation in dissected MTALs. The Akt activation was eliminated by a PI3K inhibitor and in MTALs from TLR4−/−or Toll/IL-1 receptor domain-containing adaptor-inducing IFN-β (TRIF)−/−mice. The effect of MPLA to prevent LPS inhibition of [Formula: see text] absorption also was TRIF dependent. Pretreatment with MPLA prevented LPS-induced ERK activation; this effect was dependent on PI3K. MPLA alone had no effect on [Formula: see text] absorption, and MPLA pretreatment did not prevent ERK-mediated inhibition of [Formula: see text] absorption by aldosterone, consistent with MPLA's low toxicity profile. These results demonstrate that pretreatment with MPLA prevents the effect of LPS to inhibit [Formula: see text] absorption in the MTAL. This protective effect is mediated directly through MPLA stimulation of a TLR4-TRIF-PI3K-Akt pathway that prevents LPS-induced ERK activation. These studies identify detoxified TLR4-based immunomodulators as novel potential therapeutic agents to prevent or treat renal tubule dysfunction in response to bacterial infections.

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Chlamydial infection and disease in the koala

Microbiology Australia ◽

10.1071/ma05065 ◽

2005 ◽

Vol 26 (2) ◽

pp. 65 ◽

Cited By ~ 3

Author(s):

Peter Timms

Keyword(s):

Chlamydia Trachomatis ◽

Chlamydial Infection ◽

Developmental Cycle ◽

Elementary Body ◽

Obligate Intracellular ◽

Molecular Approaches ◽

Reticulate Body ◽

Wide Range ◽

The Family ◽

Serious Disease

Chlamydiae are obligate intracellular bacterial pathogens able to infect and cause serious disease in humans, birds and a remarkably wide range of warm and cold-blooded animals. The family Chlamydiaciae have traditionally been defined by their unique biphasic developmental cycle, involving the interconversion between an extracellular survival form, the elementary body and an intracellular replicative form, the reticulate body. However, as with many other bacteria, molecular approaches including 16SrRNA sequence are becoming the standard of choice. As a consequence, the chlamydiae are in a taxonomic state of flux. Prior to 1999, the family Chlamydiaceae consisted of one genus, Chlamydia, and four species, Chlamydia trachomatis, C. psittaci, C. pecorum and C. pneumoniae. In 1999, Everett et al proposed a reclassification of Chlamydia into two genera (Chlamydia and Chlamydophila) and nine species (Chlamydia trachomatis, C. suis, and C. muridarum and Chlamydophila psittaci, C. pneumoniae, C. felis, C. pecorum, C. abortus, and C. caviae). While some of these species are thought to be host specific (C. suis ? pigs, C. muridarum ? mice, C. felis ? cats, C. caviae ? guinea pigs) many are known to infect and cause disease in a wide range of hosts.

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Host range of obligate intracellular bacterium Parachlamydia acanthamoebae

Microbiology and Immunology ◽

10.1111/j.1348-0421.2010.00265.x ◽

2010 ◽

Vol 54 (11) ◽

pp. 707-713 ◽

Cited By ~ 11

Author(s):

Yasuhiro Hayashi ◽

Shinji Nakamura ◽

Junji Matsuo ◽

Tatsuya Fukumoto ◽

Mitsutaka Yoshida ◽

...

Keyword(s):

Host Range ◽

Intracellular Bacterium ◽

Obligate Intracellular Bacterium ◽

Obligate Intracellular

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oxLDL induces endothelial cell proliferation via Rho/ROCK/Akt/p27kip1 signaling: opposite effects of oxLDL and cholesterol loading

AJP Cell Physiology ◽

10.1152/ajpcell.00249.2016 ◽

2017 ◽

Vol 313 (3) ◽

pp. C340-C351 ◽

Cited By ~ 11

Author(s):

Chongxu Zhang ◽

Crystal Adamos ◽

Myung-Jin Oh ◽

Jugajyoti Baruah ◽

Manuela A. A. Ayee ◽

...

Keyword(s):

Cell Proliferation ◽

Kinase Inhibitor ◽

Oxidized Ldl ◽

Endothelial Cell Proliferation ◽

Dependent Manner ◽

Akt Inhibitor ◽

Aortic Endothelial Cells ◽

Akt Phosphorylation ◽

Downstream Target ◽

Underlying Mechanisms

Oxidized modifications of LDL (oxLDL) play a key role in the development of endothelial dysfunction and atherosclerosis. However, the underlying mechanisms of oxLDL-mediated cellular behavior are not completely understood. Here, we compared the effects of two major types of oxLDL, copper-oxidized LDL (Cu2+-oxLDL) and lipoxygenase-oxidized LDL (LPO-oxLDL), on proliferation of human aortic endothelial cells (HAECs). Cu2+-oxLDL enhanced HAECs’ proliferation in a dose- and degree of oxidation-dependent manner. Similarly, LPO-oxLDL also enhanced HAEC proliferation. Mechanistically, both Cu2+-oxLDL and LPO-oxLDL enhance HAEC proliferation via activation of Rho, Akt phosphorylation, and a decrease in the expression of cyclin-dependent kinase inhibitor 1B (p27kip1). Both Cu2+-oxLDL or LPO-oxLDL significantly increased Akt phosphorylation, whereas an Akt inhibitor, MK2206, blocked oxLDL-induced increase in HAEC proliferation. Blocking Rho with C3 or its downstream target ROCK with Y27632 significantly inhibited oxLDL-induced Akt phosphorylation and proliferation mediated by both Cu2+- and LPO-oxLDL. Activation of RhoA was blocked by Rho-GDI-1, which also abrogated oxLDL-induced Akt phosphorylation and HAEC proliferation. In contrast, blocking Rac1 in these cells had no effect on oxLDL-induced Akt phosphorylation or cell proliferation. Moreover, oxLDL-induced Rho/Akt signaling downregulated cell cycle inhibitor p27kip1. Preloading these cells with cholesterol, however, prevented oxLDL-induced Akt phosphorylation and HAEC proliferation. These findings provide a new understanding of the effects of oxLDL on endothelial proliferation, which is essential for developing new treatments against neovascularization and progression of atherosclerosis.

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