scholarly journals The study of the remyelinating effect of leukemia-inhibitor factor and melatonin on the toxic cuprizone model of demyelination of murine cerebellar cells culture in vitro

2018 ◽  
Vol 6 (2) ◽  
Author(s):  
A. Rodnichenko ◽  
I. Labunets
Keyword(s):  
Cell Culture ◽  
Histochemical Staining ◽  
Immunocytochemical Staining ◽  
Human Leukemia ◽  
Sudan Black B ◽  
Cerebellar Cell ◽  
Leukemia Inhibitor Factor ◽  
Cerebellar Cells

The cuprizone model of toxic demyelination in vitro is widely used to study the of de- and remyelination in the CNS, as well as to address the issues of finding potential compounds that affect myelination of neuron axons.The aim of the study was to investigate the role of recombinant human leukemia inhibitory factor (rhLIF) and melatonin in remyelination, using the cuprizone demyelination model in vitro.Methods. To study the features of the demyelination and remyelination processes of neuronal axons, the culture of dissociated cerebellar cell culture of the 7-day-old FVB/N lineage mice was used. To detect the myelin sheaths, a histochemical staining with a Sudan Black B was used. To identify oligodendrocytes, immunocytochemical staining of 28-30-old-day cerebellar cells cultures for oligodendrocytes marker Olig2 was performed.Results. The direct effect of the demyelinating factor of cuprizone and remyelination agents (rhLIF and melatonin) on oligodendrocytes in vitro was confirmed. The remyelinating effect of LIF and melatonin on the restoration of myelination processes in dissociated cerebellar cell culture using histochemical and immunocytochemical staining has been revealed.Conclusions. Cuprizone-induced demyelination in vitro is associated with the death of Olig2+ oligodendrocytes and loss of myelin formation. rhLIF and melatonin are prevented the loss of oligodendrocytes and, consequently, reduced the destruction of myelin membranes.

scholarly journals Metal Complexes Derived of Diosmin with Biological Activities in vitro

2019 ◽  
Vol 9 (1) ◽  
pp. 10
Author(s):  
Maéli M. F. Civa ◽  
Dirceu G. de Souza ◽  
Renata G. Silva ◽  
Dayany da S. A. Maciel ◽  
Ricardo L. Tranquilin ◽  
...  
Keyword(s):  
Metal Complexes ◽  
Biological Activities ◽  
Coordination Complexes ◽  
Human Leukemia ◽  
Human Leukemia Cells ◽  
Ft Ir ◽  
Xrd Techniques ◽  
Better Than

The coordination of metal ions with flavonoids is applied to improve its pharmacological properties. To evaluate the role of ions on diosmin new complexes with Fe(II), Cu(II) and Co(II) ions were synthetized and characterized by UV, FT-IR and XRD techniques and surface morphology by SEM. The biological activity of coordination complexes in vitro, the antioxidant (ABTS), antibacterial (disc diffusion and MIC) and antitumoral activities (MTT) were analyzed. Diosmin when reacting with Fe(II) at 50ºC loses the sugar molecule becoming diosmetin (D) coordinated at 1D:1Fe ratio. In presence of Cu(II) and Co(II) at the same conditions besides losing the sugar, diosmin loses the methyl group at C4’ and H at C3’, producing a new ligand and complexes at 1D:2Cu or Co ratio, to produce DCu and DCo, respectively. The coordination of Cu and Fe improve the antioxidant activity of diosmin. DCo was the only presented antibacterial activity. Additionally, a specific antitumor effect of diosmin and metal complexes upon human leukemia cells was demonstrated, suggesting an immune regulatory action. The anti-melanoma activity of DCo is 10 times better than diosmin. Metal coordination could be used to improve drug activity and to give direction to a new possibility of clinical use for diosmin.

scholarly journals Tulane Virus Recognizes the A Type 3 and B Histo-Blood Group Antigens

2014 ◽  
Vol 89 (2) ◽  
pp. 1419-1427 ◽  
Author(s):  
Dongsheng Zhang ◽  
Pengwei Huang ◽  
Lu Zou ◽  
Todd L. Lowary ◽  
Ming Tan ◽  
...  
Keyword(s):  
Cell Culture ◽  
Blood Group ◽  
Culture Method ◽  
Blood Group Antigens ◽  
Content Type ◽  
Tulane Virus ◽  
A Cell ◽  
Type 3

ABSTRACTTulane virus (TV), the prototype of theRecovirusgenus in the calicivirus family, was isolated from the stools of rhesus monkeys and can be cultivatedin vitroin monkey kidney cells. TV is genetically closely related to the genusNorovirusand recognizes the histo-blood group antigens (HBGAs), similarly to human noroviruses (NoVs), making it a valuable surrogate for human NoVs. However, the precise structures of HBGAs recognized by TV remain elusive. In this study, we performed binding and blocking experiments on TV with extended HBGA types and showed that, while TV binds all four types (types 1 to 4) of the B antigens, it recognizes only the A type 3 antigen among four types of A antigens tested. The requirements for HBGAs in TV replication were demonstrated by blocking of TV replication in cell culture using the A type 3/4 and B saliva samples. Similar results were also observed in oligosaccharide-based blocking assays. Importantly, the previously reported, unexplained increase in TV replication by oligosaccharide in cell-based blocking assays has been clarified, which will facilitate the application of TV as a surrogate for human NoVs.IMPORTANCEOur understanding of the role of HBGAs in NoV infection has been significantly advanced in the past decade, but direct evidence for HBGAs as receptors for human NoVs remains lacking due to a lack of a cell culture method. TV recognizes HBGAs and can replicatein vitro, providing a valuable surrogate for human NoVs. However, TV binds to some but not all saliva samples from A-positive individuals, and an unexplained observation of synthetic oligosaccharide blocking of TV binding has been reported. These issues have been resolved in this study.

scholarly journals Modeling hepatitis C virus kinetics during liver transplantation highlights the role of the liver in virus clearance

2020 ◽  
Author(s):  
Louis Shekhtman ◽  
Miquel Navasa ◽  
Natasha Sansone ◽  
Gonzalo Crespo ◽  
Gitanjali Subramanya ◽  
...  
Keyword(s):  
Cell Culture ◽  
Liver Transplantation ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Clearance Rate ◽  
Culture Medium ◽  
Viral Clearance ◽  
Huh7 Cells

AbstractWhile the liver, specifically hepatocytes, are widely accepted as the main source for hepatitis C virus (HCV) production, the role of the liver/hepatocytes in the clearance of circulating HCV remains largely unknown. Here we evaluated the function of the liver/hepatocytes in clearing virus from the circulation by investigating viral clearance during liver transplantation and from culture medium in vitro. Frequent HCV kinetic data during liver transplantation were recorded from 5 individuals throughout the anhepatic (AH) phase and for 4 hours after reperfusion (RP), along with recordings of fluid balances. Using mathematical modeling, the serum viral clearance rate, c, was estimated. Analogously, we monitored the clearance rate of HCV at 37°C from culture medium in vitro in the absence and presence of chronically infected Huh7 human hepatoma cells. During the AH phase, in 3 transplant cases viral levels remained at pre-AH levels, while in the other 2 cases HCV declined (half-life, t1/2~1h). Immediately post-RP, virus declined in a biphasic manner in Cases 1-4 consisting of an extremely rapid (median t1/2=5min) decline followed by a slower decline (HCV t1/2=67min). In Case 5, HCV remained at the same level post-RP as at the end of AH. Declines in virus level were not explained by adjusting for dilution from IV fluid and blood products. Consistent with what was observed in the majority of patients in the anhepatic phase, the t1/2 of HCV in cell culture was much longer in the absence of chronically HCV-infected Huh7 cells. Therefore, kinetic and modeling results from both in vivo liver transplantation cases and in vitro cell culture studies suggest that the liver plays a major role in clearing HCV from the circulation.

Granular cells are required for encapsulation of foreign targets by insect haemocytes

1996 ◽  
Vol 109 (8) ◽  
pp. 2053-2060 ◽  
Author(s):  
L.L. Pech ◽  
M.R. Strand
Keyword(s):  
Cell Adhesion ◽  
Granular Cell ◽  
Immunocytochemical Staining ◽  
Granular Cells ◽  
Recognition Sequence ◽  
Capsule Formation ◽  
Insect Haemocytes ◽  
Dependent Cell

Haemocytes play an essential role in defending invertebrates against pathogens and parasites that enter their haemocoel. A primary defense response is encapsulation; a process in which haemocytes attach to the foreign organism and kill it. Whether encapsulation requires cooperation between specific subpopulations of haemocytes is unknown. Using purified subpopulations of haemocytes and an in vitro encapsulation assay, we investigated the process of capsule formation in the insect Pseudoplusia includens. Immunocytochemical staining revealed that capsule formation involves a three step process. Encapsulation began when granular cells attached to the foreign target. This was followed by attachment of multiple layers of plasmatocytes. Termination of capsule formation occurred when a subpopulation of granular cells formed a monolayer around the periphery of the capsule. Neither granular cells nor plasmatocytes were capable of forming a capsule independently. However, plasmatocytes encapsulated targets if granular cells were present or if targets were preincubated in medium conditioned by granular cells. The effect of granular cell-conditioned medium could be blocked by the addition of the cell adhesion recognition sequence, RGDS, but not by RGES. These results demonstrate experimentally that granular cells are required for encapsulation of foreign targets by plasmatocytes in vitro, and that the role of granular cells in this process involves an RGD-dependent cell adhesion mechanism.

scholarly journals Human Th17 cells produce a soluble mediator that increases podocyte motility via signaling pathways that mimic PAR-1 activation

2019 ◽  
Vol 317 (4) ◽  
pp. F913-F921 ◽  
Author(s):  
Carl J. May ◽  
Gavin I. Welsh ◽  
Musleeha Chesor ◽  
Phillipa J. Lait ◽  
Lauren P. Schewitz-Bowers ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Cultures ◽  
Th17 Cell ◽  
Th17 Cells ◽  
Culture Supernatant ◽  
Cell Culture Supernatant ◽  
Soluble Mediator ◽  
Stimulation Of

The specific pathogenesis of idiopathic nephrotic syndrome (NS) is poorly understood, and the role of immune mediators remains contentious. However, there is good evidence for the role of a circulating factor, and we recently postulated circulating proteases as candidate factors. Immunosuppressive therapy with glucocorticoids (GCs) and T cell inhibitors are widely used in the clinical treatment of NS. Given that T helper (CD4+) cells expressing IL-17A (so-called Th17 cells) have recently been reported to be resistant to GC treatment, and GC resistance remains a major challenge in the management of NS, we hypothesized that Th17 cells produce a circulating factor that is capable of signaling to the podocyte and inducing deleterious phenotypic changes. To test this, we generated human Th17 cells from healthy volunteers and added the supernatants from these T cell cultures to conditionally immortalized human podocytes in vitro. This demonstrated that podocytes treated with Th17 cell culture supernatant, as well as with patient disease plasma, showed significant stimulation of JNK and p38 MAPK pathways and an increase in motility, which was blocked using a JNK inhibitor. We have previously shown that nephrotic plasma elicits a podocyte response via protease-activated receptor-1 (PAR-1). Stimulation of PAR-1 in podocytes elicited the same signaling response as Th17 cell culture supernatant treatment. Equally, protease inhibitors with Th17 cell culture treatment blocked the signaling response. This was not replicated by the reagents added to Th17 cell cultures or by IL-17A. Hence, we conclude that an undefined soluble mediator produced by Th17 cells mimics the deleterious effect of PAR-1 activation in vitro. Given the association between pathogenic subsets of Th17 cells and GC resistance, these observations have potential therapeutic relevance for patients with NS.

scholarly journals Strand-Specific RNA Synthesis Determinants in the RNA-Dependent RNA Polymerase of Poliovirus

2004 ◽  
Vol 78 (9) ◽  
pp. 4397-4407 ◽  
Author(s):  
Christopher T. Cornell ◽  
Rushika Perera ◽  
Jo Ellen Brunner ◽  
Bert L. Semler
Keyword(s):  
Cell Culture ◽  
Rna Polymerase ◽  
Rna Synthesis ◽  
Rna Replication ◽  
Amino Acid Sequences ◽  
Rna Dependent Rna Polymerase ◽  
Amino Terminal ◽  
Carboxy Terminal

ABSTRACT The viral RNA-dependent RNA polymerase (3Dpol) is highly conserved between the closely related enteroviruses poliovirus type 1 (PV1) and coxsackievirus B3 (CVB3). In this study, we generated PV1/CVB3 chimeric polymerase sequences in the context of full-length poliovirus transcripts to determine the role of different subdomains within the RNA-dependent RNA polymerase of PV1 that are required for functions critical for RNA replication in vitro and in cell culture. The substitution of CVB3 sequences in the carboxy-terminal portion (thumb subdomain) of the polymerase resulted in transcripts incapable of RNA replication. In contrast, three of the seven chimeras were capable of synthesizing RNA, albeit to reduced levels compared to that of wild-type PV1 RNA. Interestingly, one of the replication-competent chimeras (CPP) displayed an inability to generate positive strands, indicating the presence of amino-terminal sequences within the 3D polymerase and/or the 3D domain of the 3CD precursor polypeptide that are necessary for the assembly of strand-specific RNA synthesis complexes. In some constructs, the partial reestablishment of PV1 amino acid sequences in this region was capable of rescuing RNA replication in vitro and in cell culture.

scholarly journals Enhanced BDNF Actions Following Acute Hypoxia Facilitate HIF-1α-Dependent Upregulation of Cav3-T-Type Ca2+ Channels in Rat Cardiomyocytes

Membranes ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 470
Author(s):  
Masaki Morishima ◽  
Takafumi Fujita ◽  
Satoshi Osagawa ◽  
Hiroshi Kubota ◽  
Katsushige Ono
Keyword(s):  
Acute Hypoxia ◽  
Hypoxia Inducible Factor ◽  
Immunocytochemical Staining ◽  
Rat Cardiomyocytes ◽  
Neonatal Cardiomyocytes ◽  
Artery Disease ◽  
Interfering Rna ◽  
Ca2 Channels

Brain-derived neurotrophic factor (BDNF) has recently been recognized as a cardiovascular regulator particularly in the diseased condition, including coronary artery disease, heart failure, cardiomyopathy, and hypertension. Here, we investigate the role of BDNF on the T-type Ca2+ channel, Cav3.1 and Cav3.2, in rat neonatal cardiomyocytes exposed to normoxia (21% O2) and acute hypoxia (1% O2) in vitro for up to 3 h. The exposure of cardiomyocytes to hypoxia (1 h, 3 h) caused a significant upregulation of the mRNAs for hypoxia-inducible factor 1α (Hif1α), Cav3.1, Cav3.2 and Bdnf, but not tropomyosin-related kinase receptor B (TrkB). The upregulation of Cav3.1 and Cav3.2 caused by hypoxia was completely halted by small interfering RNA (siRNA) targeting Hif1a (Hif1a-siRNA) or Bdnf (Bdnf-siRNA). Immunocytochemical staining data revealed a distinct upregulation of Cav3.1- and Cav3.2-proteins caused by hypoxia in cardiomyocytes, which was markedly suppressed by Bdnf-siRNA. These results unveiled a novel regulatory action of BDNF on the T-type Ca2+ channels expression through the HIF-1α-dependent pathway in cardiomyocytes.

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