scholarly journals Sensitivity of Rodent Microglia to Kynurenines in Models of Epilepsy and Inflammation In Vivo and In Vitro: Microglia Activation Is Inhibited by Kynurenic Acid and the Synthetic Analogue SZR104

2020 ◽  
Vol 21 (23) ◽  
pp. 9333
Author(s):  
Noémi Lajkó ◽  
Diana Kata ◽  
Melinda Szabó ◽  
Adrienne Mátyás ◽  
Karolina Dulka ◽  
...  
Keyword(s):  
Calcium Binding ◽  
Kynurenic Acid ◽  
Synthetic Analogue ◽  
Endogenous Modulator ◽  
Genes Encoding ◽  
Immunosuppressive Action ◽  
Kynurenic Acid Analogue ◽  
Phagocytotic Activity

Kynurenic acid is an endogenous modulator of ionotropic glutamate receptors and a suppressor of the immune system. Since glutamate and microglia are important in the pathogenesis of epilepsy, we investigated the possible action of the synthetic kynurenic acid analogue, SZR104, in epileptic mice and the action of kynurenic acid and SZR104 on the phagocytotic activity of cultured microglia cells. Pilocarpine epilepsy was used to test the effects of SZR104 on morphological microglia transformation, as evaluated through ionized calcium-binding adaptor molecule 1 (Iba1) immunohistochemistry. Microglia-enriched rat secondary cultures were used to investigate phagocytosis of fluorescent microbeads and Iba1 protein synthesis in control and lipopolysaccharide-challenged cultures. SZR104 inhibited microglia transformation following status epilepticus. Kynurenic acid and SZR104 inhibited lipopolysaccharide-stimulated phagocytotic activity of microglia cells. Although kynurenic acid and its analogues proved to be glutamate receptor antagonists, their immunosuppressive action was dominant in epilepsy. The inhibition of phagocytosis in vitro raised the possibility of the inhibition of genes encoding inflammatory cytokines in microglial cells.

scholarly journals In vivo transcriptome analysis provides insights into host-dependent expression of virulence factors by Yersinia entomophaga MH96, during infection of Galleria mellonella

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Amber R Paulson ◽  
Maureen O’Callaghan ◽  
Xue-Xian Zhang ◽  
Paul B Rainey ◽  
Mark R H Hurst
Keyword(s):  
Galleria Mellonella ◽  
Functional Enrichment ◽  
Natural Environments ◽  
Insecticidal Toxin ◽  
Heat Stable ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Cell Densities

Abstract The function of microbes can be inferred from knowledge of genes specifically expressed in natural environments. Here, we report the in vivo transcriptome of the entomopathogenic bacterium Yersinia entomophaga MH96, captured during initial, septicemic, and pre-cadaveric stages of intrahemocoelic infection in Galleria mellonella. A total of 1285 genes were significantly upregulated by MH96 during infection; 829 genes responded to in vivo conditions during at least one stage of infection, 289 responded during two stages of infection, and 167 transcripts responded throughout all three stages of infection compared to in vitro conditions at equivalent cell densities. Genes upregulated during the earliest infection stage included components of the insecticidal toxin complex Yen-TC (chi1, chi2, and yenC1), genes for rearrangement hotspot element containing protein yenC3, cytolethal distending toxin cdtAB, and vegetative insecticidal toxin vip2. Genes more highly expressed throughout the infection cycle included the putative heat-stable enterotoxin yenT and three adhesins (usher-chaperone fimbria, filamentous hemagglutinin, and an AidA-like secreted adhesin). Clustering and functional enrichment of gene expression data also revealed expression of genes encoding type III and VI secretion system-associated effectors. Together these data provide insight into the pathobiology of MH96 and serve as an important resource supporting efforts to identify novel insecticidal agents.

scholarly journals Kynurenic Acid Accelerates Healing of Corneal Epithelium In Vitro and In Vivo

2021 ◽  
Vol 14 (8) ◽  
pp. 753
Author(s):  
Anna Matysik-Woźniak ◽  
Waldemar A. Turski ◽  
Monika Turska ◽  
Roman Paduch ◽  
Mirosław Łańcut ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Corneal Epithelium ◽  
Kynurenic Acid ◽  
Pharmaceutical Products ◽  
Corneal Epithelial Cell ◽  
Eye Drops ◽  
Conjunctival Epithelium ◽  
Corneal Erosion

Kynurenic acid (KYNA) is an endogenous compound with a multidirectional effect. It possesses antiapoptotic, anti-inflammatory, and antioxidative properties that may be beneficial in the treatment of corneal injuries. Moreover, KYNA has been used successfully to improve the healing outcome of skin wounds. The aim of the present study is to evaluate the effects of KYNA on corneal and conjunctival cells in vitro and the re-epithelization of corneal erosion in rabbits in vivo. Normal human corneal epithelial cell (10.014 pRSV-T) and conjunctival epithelial cell (HC0597) lines were used. Cellular metabolism, cell viability, transwell migration, and the secretion of IL-1β, IL-6, and IL-10 were determined. In rabbits, after corneal de-epithelization, eye drops containing 0.002% and 1% KYNA were applied five times a day until full recovery. KYNA decreased metabolism but did not affect the proliferation of the corneal epithelium. It decreased both the metabolism and proliferation of conjunctival epithelium. KYNA enhanced the migration of corneal but not conjunctival epithelial cells. KYNA reduced the secretion of IL-1β and IL-6 from the corneal epithelium, leaving IL-10 secretion unaffected. The release of all studied cytokines from the conjunctival epithelium exposed to KYNA was unchanged. KYNA at higher concentration accelerated the healing of the corneal epithelium. These favorable properties of KYNA suggest that KYNA containing topical pharmaceutical products can be used in the treatment of ocular surface diseases.

scholarly journals Designing P. aeruginosa synthetic phages with reduced genomes

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Diana P. Pires ◽  
Rodrigo Monteiro ◽  
Dalila Mil-Homens ◽  
Arsénio Fialho ◽  
Timothy K. Lu ◽  
...  
Keyword(s):  
Galleria Mellonella ◽  
Antibacterial Properties ◽  
Genetically Engineered ◽  
In Vivo Studies ◽  
Infection Model ◽  
Promising Therapeutic Approach ◽  
Genes Encoding ◽  
First Time

AbstractIn the era where antibiotic resistance is considered one of the major worldwide concerns, bacteriophages have emerged as a promising therapeutic approach to deal with this problem. Genetically engineered bacteriophages can enable enhanced anti-bacterial functionalities, but require cloning additional genes into the phage genomes, which might be challenging due to the DNA encapsulation capacity of a phage. To tackle this issue, we designed and assembled for the first time synthetic phages with smaller genomes by knocking out up to 48% of the genes encoding hypothetical proteins from the genome of the newly isolated Pseudomonas aeruginosa phage vB_PaeP_PE3. The antibacterial efficacy of the wild-type and the synthetic phages was assessed in vitro as well as in vivo using a Galleria mellonella infection model. Overall, both in vitro and in vivo studies revealed that the knock-outs made in phage genome do not impair the antibacterial properties of the synthetic phages, indicating that this could be a good strategy to clear space from phage genomes in order to enable the introduction of other genes of interest that can potentiate the future treatment of P. aeruginosa infections.

scholarly journals Exosomal S100A4 derived from highly metastatic hepatocellular carcinoma cells promotes metastasis by activating STAT3

2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Haoting Sun ◽  
Chaoqun Wang ◽  
Beiyuan Hu ◽  
Xiaomei Gao ◽  
Tiantian Zou ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cancer Progression ◽  
Calcium Binding ◽  
Tumor Metastasis ◽  
Metastatic Potential ◽  
Functional Roles ◽  
Stat3 Phosphorylation ◽  
Hcc Cells

AbstractIntercellular cross-talk plays important roles in cancer progression and metastasis. Yet how these cancer cells interact with each other is still largely unknown. Exosomes released by tumor cells have been proved to be effective cell-to-cell signal mediators. We explored the functional roles of exosomes in metastasis and the potential prognostic values for hepatocellular carcinoma (HCC). Exosomes were extracted from HCC cells of different metastatic potentials. The metastatic effects of exosomes derived from highly metastatic HCC cells (HMH) were evaluated both in vitro and in vivo. Exosomal proteins were identified with iTRAQ mass spectrum and verified in cell lines, xenograft tumor samples, and functional analyses. Exosomes released by HMH significantly enhanced the in vitro invasion and in vivo metastasis of low metastatic HCC cells (LMH). S100 calcium-binding protein A4 (S100A4) was identified as a functional factor in exosomes derived from HMH. S100A4rich exosomes significantly promoted tumor metastasis both in vitro and in vivo compared with S100A4low exosomes or controls. Moreover, exosomal S100A4 could induce expression of osteopontin (OPN), along with other tumor metastasis/stemness-related genes. Exosomal S100A4 activated OPN transcription via STAT3 phosphorylation. HCC patients with high exosomal S100A4 in plasma also had a poorer prognosis. In conclusion, exosomes from HMH could promote the metastatic potential of LMH, and exosomal S100A4 is a key enhancer for HCC metastasis, activating STAT3 phosphorylation and up-regulating OPN expression. This suggested exosomal S100A4 to be a novel prognostic marker and therapeutic target for HCC metastasis.

scholarly journals Neuroprotective Effect of Optimized Yinxieling Formula in 6-OHDA-Induced Chronic Model of Parkinson’s Disease through the Inflammation Pathway

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Renrong Wei ◽  
Cuiping Rong ◽  
Qingfeng Xie ◽  
Shouhai Wu ◽  
Yuchao Feng ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Calcium Binding ◽  
Dopaminergic Neurons ◽  
Neuroprotective Effect ◽  
Interleukin 1 ◽  
Mrna Levels ◽  
Cox 2

Parkinson’s disease (PD) is characterized by progressive degeneration of dopaminergic neurons in the substantia nigra (SN)-striatum circuit, which is associated with glial activation and consequent chronic neuroinflammation. Optimized Yinxieling Formula (OYF) is a Chinese medicine that exerts therapeutical effect and antiinflammation property on psoriasis. Our previous study has proven that pretreatment with OYF could regulate glia-mediated inflammation in an acute mouse model of PD induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. Given that PD is a chronic degeneration disorder, this study applied another PD animal model induced by striatal injection of 6-hydroxydopamine (6-OHDA) to mimic the progressive damage of the SN-striatum dopamine system in rats. The OYF was administrated in the manner of pretreatment plus treatment. The effects of the OYF on motor behaviors were assessed with the apomorphine-induced rotation test and adjusting steps test. To confirm the effect of OYF on dopaminergic neurons and glia activation in this model, we analyzed the expression of tyrosine hydroxylase (TH) and glia markers, ionized calcium-binding adapter molecule 1 (Iba-1), and glial fibrillary acidic protein (GFAP) in the SN region of the rat PD model. Inflammation-associated factors, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2), were further evaluated in this model and in interferon-γ- (INF-γ-) induced murine macrophages RAW264.7 cells. The results from the in vivo study showed that OYF reversed the motor behavioral dysfunction in 6-OHDA-induced PD rats, upregulated the TH expression, decreased the immunoreactivity of Iba-1 and GFAP, and downregulated the mRNA levels of TNF-α and COX-2. The OYF also trended to decrease the mRNA levels of IL-1β and iNOS in vivo. The results from the in vitro study showed that OYF significantly decreased the mRNA levels of TNF-α, IL-1β, IL-6, iNOS, and COX-2. Therefore, this study suggests that OYF exerts antiinflammatory effects, which might be related to the protection of dopaminergic neurons in 6-OHDA-induced chronic neurotoxicity.

scholarly journals Microtubule Actin Cross-Linking Factor (Macf)

1999 ◽  
Vol 147 (6) ◽  
pp. 1275-1286 ◽  
Author(s):  
Conrad L. Leung ◽  
Dongming Sun ◽  
Min Zheng ◽  
David R. Knowles ◽  
Ronald K.H. Liem
Keyword(s):  
Calcium Binding ◽  
Coiled Coil ◽  
Specific Protein ◽  
Binding Domain ◽  
Full Length ◽  
Actin Binding ◽  
Cross Linking ◽  
Actin Binding Domain

We cloned and characterized a full-length cDNA of mouse actin cross-linking family 7 (mACF7) by sequential rapid amplification of cDNA ends–PCR. The completed mACF7 cDNA is 17 kb and codes for a 608-kD protein. The closest relative of mACF7 is the Drosophila protein Kakapo, which shares similar architecture with mACF7. mACF7 contains a putative actin-binding domain and a plakin-like domain that are highly homologous to dystonin (BPAG1-n) at its NH2 terminus. However, unlike dystonin, mACF7 does not contain a coiled–coil rod domain; instead, the rod domain of mACF7 is made up of 23 dystrophin-like spectrin repeats. At its COOH terminus, mACF7 contains two putative EF-hand calcium-binding motifs and a segment homologous to the growth arrest–specific protein, Gas2. In this paper, we demonstrate that the NH2-terminal actin-binding domain of mACF7 is functional both in vivo and in vitro. More importantly, we found that the COOH-terminal domain of mACF7 interacts with and stabilizes microtubules. In transfected cells full-length mACF7 can associate not only with actin but also with microtubules. Hence, we suggest a modified name: MACF (microtubule actin cross-linking factor). The properties of MACF are consistent with the observation that mutations in kakapo cause disorganization of microtubules in epidermal muscle attachment cells and some sensory neurons.

Effect of steroids on cartilage metabolism "in vitro"

1986 ◽  
Vol 113 (4_Suppl) ◽  
pp. S35-S40 ◽  
Author(s):  
Marc V.L. DU CAJU ◽  
Raoul P. ROOMAN
Keyword(s):  
Growth Factor ◽  
Thymidine Incorporation ◽  
Normal Serum ◽  
Synthetic Analogue ◽  
Factor Activity ◽  
Poor Growth ◽  
Cartilage Metabolism ◽  
Glucocorticoid Administration

ABSTRACT Conditions characterized by high levels of glucocorticoids are associated with poor growth. Serum somatomedin or insulin-like growth factor activity measured by cartilage bioassay systems is low, but is generally not accompanied by a fall in somatomedin concentration. Hydrocortisone and a synthetic analogue, dexamethasone, impaired the serum stimulated "in vitro" 35S sulphate and 3H-thymidine incorporation in porcine rib cartilage at physiological concentrations. Hydrocortisone added at a concentration of 0,1 μg/ml decreased the potency of normal serum to 50 % of controls. Dexamethasone was at least 10 times more potent. Removal of "in vitro" or "in vivo" administered hydrocortisone with dextran-coated charcoal restored the sulphate and thymidine activity to normal. We conclude that physiological amounts of glucocorticoids inhibit the "in vitro" porcine cartilage metabolism. Glucocorticoid administration "in vivo" does not abolish the activity of the cartilage stimulating effect of serum but affects cartilage metabolism directly or by the induction of locally produced inhibitors of cartilage metabolism.

scholarly journals Virulent Shigella flexneri subverts the host innate immune response through manipulation of antimicrobial peptide gene expression

2008 ◽  
Vol 205 (5) ◽  
pp. 1121-1132 ◽  
Author(s):  
Brice Sperandio ◽  
Béatrice Regnault ◽  
Jianhua Guo ◽  
Zhi Zhang ◽  
Samuel L. Stanley ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Shigella Flexneri ◽  
Host Cells ◽  
Virulence Plasmid ◽  
Cationic Peptides ◽  
Immunity Genes ◽  
Genes Encoding ◽  
Peptide Gene

Antimicrobial factors are efficient defense components of the innate immunity, playing a crucial role in the intestinal homeostasis and protection against pathogens. In this study, we report that upon infection of polarized human intestinal cells in vitro, virulent Shigella flexneri suppress transcription of several genes encoding antimicrobial cationic peptides, particularly the human β-defensin hBD-3, which we show to be especially active against S. flexneri. This is an example of targeted survival strategy. We also identify the MxiE bacterial regulator, which controls a regulon encompassing a set of virulence plasmid-encoded effectors injected into host cells and regulating innate signaling, as being responsible for this dedicated regulatory process. In vivo, in a model of human intestinal xenotransplant, we confirm at the transcriptional and translational level, the presence of a dedicated MxiE-dependent system allowing S. flexneri to suppress expression of antimicrobial cationic peptides and promoting its deeper progression toward intestinal crypts. We demonstrate that this system is also able to down-regulate additional innate immunity genes, such as the chemokine CCL20 gene, leading to compromised recruitment of dendritic cells to the lamina propria of infected tissues. Thus, S. flexneri has developed a dedicated strategy to weaken the innate immunity to manage its survival and colonization ability in the intestine.

scholarly journals Reconstitution of the Human tRNA Splicing Endonuclease Complex: insight into the regulation of pre-tRNA cleavage

2019 ◽  
Author(s):  
Cassandra K. Hayne ◽  
Casey A. Schmidt ◽  
A. Gregory Matera ◽  
Robin E. Stanley
Keyword(s):  
Animal Cells ◽  
Trna Splicing ◽  
Polynucleotide Kinase ◽  
Genes Encoding ◽  
Intron Removal ◽  
And Function ◽  
Insight Into ◽  
Negative Modulator

ABSTRACTThe splicing of tRNA introns is a critical step in pre-tRNA maturation. In archaea and eukaryotes, tRNA intron removal is catalyzed by the tRNA splicing endonuclease (TSEN) complex. Eukaryotic TSEN is comprised of four core subunits (TSEN54, TSEN2, TSEN34, and TSEN15). The human TSEN complex additionally co-purifies with the polynucleotide kinase CLP1; however, CLP1’s role in tRNA splicing remains unclear. Mutations in genes encoding all four TSEN subunits, as well as CLP1, are known to cause neurodegenerative disorders, yet the mechanisms underlying the pathogenesis of these disorders are unknown. Here, we developed a recombinant system that produces active TSEN complex. Co-expression of all four TSEN subunits is required for efficient formation and function of the complex. We show that human CLP1 associates with the active TSEN complex, but is not required for tRNA intron cleavage in vitro. Moreover, RNAi knockdown of the Drosophila CLP1 orthologue, cbc, promotes biogenesis of mature tRNAs and circularized tRNA introns (tricRNAs) in vivo. Collectively, these and other findings suggest that CLP1/cbc plays a regulatory role in tRNA splicing by serving as a negative modulator of the direct tRNA ligation pathway in animal cells.

Sign in / Sign up

Export Citation Format

Share Document