Reversible and Photoresponsive Immobilization of Nonadherent Cells by Spiropyran-Conjugated PEG–Lipids

2019 ◽  
Vol 2 (1) ◽  
pp. 33-38 ◽  
Author(s):  
Shin Izuta ◽  
Satoshi Yamaguchi ◽  
Takahiro Kosaka ◽  
Akimitsu Okamoto
Keyword(s):  
Nonadherent Cells

Acoustofection: High-Frequency Vibrational Membrane Permeabilization for Intracellular siRNA Delivery into Nonadherent Cells

2021 ◽  
Vol 4 (3) ◽  
pp. 2781-2789
Author(s):  
Shwathy Ramesan ◽  
Amgad R. Rezk ◽  
Paula M. Cevaal ◽  
Christina Cortez-Jugo ◽  
Jori Symons ◽  
...  
Keyword(s):  
High Frequency ◽  
Sirna Delivery ◽  
Membrane Permeabilization ◽  
Nonadherent Cells

scholarly journals Anti-CD63 antibodies suppress IgE-dependent allergic reactions in vitro and in vivo

2005 ◽  
Vol 201 (3) ◽  
pp. 385-396 ◽  
Author(s):  
Stefan Kraft ◽  
Tony Fleming ◽  
James M. Billingsley ◽  
Shih-Yao Lin ◽  
Marie-Hélène Jouvin ◽  
...  
Keyword(s):  
Pi3k Pathway ◽  
Therapeutic Agents ◽  
Allergic Reactions ◽  
Ecm Proteins ◽  
Broad Array ◽  
High Affinity Ige Receptor ◽  
Nonadherent Cells ◽  
Tetraspanin Cd63

High-affinity IgE receptor (FcεRI) cross-linking on mast cells (MCs) induces secretion of preformed allergy mediators (degranulation) and synthesis of lipid mediators and cytokines. Degranulation produces many symptoms of immediate-type allergic reactions and is modulated by adhesion to surfaces coated with specific extracellular matrix (ECM) proteins. The signals involved in this modulation are mostly unknown and their contribution to allergic reactions in vivo is unclear. Here we report the generation of monoclonal antibodies that potently suppress FcεRI-induced degranulation, but not leukotriene synthesis. We identified the antibody target as the tetraspanin CD63. Tetraspanins are membrane molecules that form multimolecular complexes with a broad array of molecules including ECM protein-binding β integrins. We found that anti-CD63 inhibits MC adhesion to fibronectin and vitronectin. Furthermore, anti-CD63 inhibits FcεRI-mediated degranulation in cells adherent to those ECM proteins but not in nonadherent cells. Thus the inhibition of degranulation by anti-CD63 correlates with its effect on adhesion. In support of a mechanistic linkage between the two types of inhibition, anti-CD63 had no effect on FcεRI-induced global tyrosine phosphorylation and calcium mobilization but impaired the Gab2–PI3K pathway that is known to be essential for both degranulation and adhesion. Finally, we showed that these antibodies inhibited FcεRI-mediated allergic reactions in vivo. These properties raise the possibility that anti-CD63 could be used as therapeutic agents in MC-dependent diseases.

scholarly journals An engraved surface induces weak adherence and high proliferation of nonadherent cells and microorganisms during culture

BioTechniques ◽  
2020 ◽  
Vol 69 (2) ◽  
pp. 113-125
Author(s):  
Sunil Thomas
Keyword(s):  
16S Rrna ◽  
Culture Medium ◽  
Petri Dish ◽  
Adherent Cells ◽  
Plastic Dish ◽  
Nonadherent Cells

When cells are cultured in a Petri dish, the adherent cells attach to the bottom of the dish; whereas, the nonadherent cells float in the culture medium. It was observed that nonadherent cells could be induced to adherent-like cells when cultured in an engraved plastic dish (biosimulator). The adherence of these cells to the engraved surface could be prevented with inhibitors specific for adhesion. It was also observed that culturing microorganisms of the environment in a biosimulator induced weak adhesion and high proliferation. Analysis of the microbiome using 16S rRNA profiling demonstrated that the biosimulator was more efficient in inducing proliferation of several phyla of microorganisms compared with culture by conventional techniques.

scholarly journals Amphotropic retrovirus vector transfer of the v-ras oncogene to human hematopoietic and stromal cells in continuous bone marrow cultures

Blood ◽  
1985 ◽  
Vol 65 (3) ◽  
pp. 744-752
Author(s):  
L Rothstein ◽  
JH Pierce ◽  
V Klassen ◽  
JS Greenberger
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
High Efficiency ◽  
Leukemia Virus ◽  
Reverse Transcriptase Activity ◽  
Pseudotype Virus ◽  
Bone Marrow Cultures ◽  
Ras P21 ◽  
Nonadherent Cells ◽  
Marrow Cultures

Human continuous bone marrow cultures were established from intraoperative marrow specimens and infected with amphotropic murine leukemia virus (Am-MuLV) pseudotypes of Kirsten or Harvey murine sarcoma virus, and the biologic effects were compared with mouse continuous bone marrow cultures. Cultures were tested for production of total nonadherent granulocytes and granulocyte-macrophage progenitor cells (GM-CFUc); virus replication by supernatant reverse transcriptase activity; percentage of adherent and nonadherent cells and GM-CFUc that released virus by infectious center assay; and for synthesis of Harvey ras p21 protein. High-efficiency, stable Am-MuLV infection of over 90% of human marrow-culture nonadherent and adherent cells and both seven- and 14-day GM-CFUc were detected as Kirsten or Harvey pseudotype virus release by infectious center assay. Synthesis of Harvey ras p21 was detected in the adherent and nonadherent cell populations of human as well as mouse continuous marrow cultures infected with Kirsten or Harvey pseudotype virus. In contrast to data with mouse cultures, cumulative production of GM-CFUc and differentiated granulocytes in human cultures was not detectably altered by Harvey or Kirsten virus infection, and all cultures ceased to produce hematopoietic cells by 20 weeks. Of 54 virus-infected cultures in ten separate experiments, 13 produced a second peak of nonadherent cells (greater than 10(5) per flask) after 20 weeks, significantly more frequently than did control uninfected cultures (one of 32). When subcultured, these harvests produced permanent Epstein-Barr virus (EBV)-transformed pre-B cell lines that released the original inoculating pseudotype virus. Thus, Am- MuLV is a potentially valuable vector for inserting genetic sequences by recombinant techniques into human hematopoietic and stromal cells in culture; however, activation of EBV may be a significant complication.

scholarly journals Lymphokine abnormalities in aplastic anemia: implications for the mechanism of action of antithymocyte globulin

Blood ◽  
1985 ◽  
Vol 65 (2) ◽  
pp. 407-413
Author(s):  
P Gascon ◽  
NC Zoumbos ◽  
G Scala ◽  
JY Djeu ◽  
JG Moore ◽  
...  
Keyword(s):  
Aplastic Anemia ◽  
Mechanism Of Action ◽  
Mononuclear Cells ◽  
Interleukin 2 ◽  
Lymphocyte Function ◽  
Normal Range ◽  
Flow Microfluorometry ◽  
Nonadherent Cells ◽  
Hematopoietic Response

Anti-thymocyte globulin (ATG) provides effective therapy for many patients with aplastic anemia, and its mechanism of action has been presumed to be secondary to lymphocytotoxicity. However, our studies of lymphocyte function in aplastic anemia show marked abnormalities of lymphokine production, which ATG may modulate. In 12 of 17 patients with aplastic anemia, interleukin 2 (IL2) production was markedly elevated in vitro (P less than .01 by paired statistical analysis). Expression of the IL2 receptor, or Tac antigen, on peripheral lymphocytes assessed by flow microfluorometry was also increased above the normal range in 11 of 15 cases. Studies of ATG suggested that it might act to stimulate lymphocyte function. In vitro, ATG is a mitogen, as measured by incorporation of 3H-thymidine into blood mononuclear cells; the response of cells to ATG from patients with aplastic anemia was exaggerated in comparison with normals. Cell proliferation was accompanied by production of IL2 to levels that were, in some cases, similar to those obtained with lectin stimulation. Finally, supernatants from lymphocytes cultured in the presence of ATG were able to replace adherent cells in providing growth factors for the support of nonadherent cells in methylcellulose hematopoietic colony assays. These results provide a mechanism for an “immunostimulatory” action of ATG in effecting hematopoietic response in some patients with aplastic anemia.

scholarly journals Antiproliferative and differentiative effects of recombinant interleukin-4 on a granulocyte colony-stimulating factor-dependent myeloblastic leukemic cell line

Blood ◽  
1991 ◽  
Vol 78 (2) ◽  
pp. 471-478
Author(s):  
Y Imai ◽  
N Nara ◽  
S Tohda ◽  
K Nagata ◽  
T Suzuki ◽  
...  
Keyword(s):  
Cell Line ◽  
Leukemic Cell ◽  
Interleukin 4 ◽  
Tnf Alpha ◽  
Adherent Cells ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Leukemic Cell Line ◽  
Nonadherent Cells ◽  
Stimulating Factor

The effect of recombinant human interleukin-4 (IL-4) on a granulocyte colony-stimulating factor (G-CSF)-dependent human myeloblastic leukemic cell line, OCI-AML1a, was investigated. IL-4 suppressed the clonogenic cell growth in methylcellulose culture, inhibited the uptake of 3H thymidine in a dose-dependent manner at 5 to 100 U/mL, and consequently suppressed the growth of clonogenic cells in short- and long-term suspension cultures. In addition, IL-4 markedly increased the number of adherent cells. These adherent cells were alpha-naphthyl-butyrate (alpha-NB) esterase-positive and showed macrophage-like appearance, increased expression of CD14, CD11b, CD23, and Ia, and significantly decreased clonogenicity. On the other hand, nonadherent cells growing in suspension showed only slight increase in proportion of alpha-NB esterase-positive or monocyte/macrophage-like cells and increased CD23 expression by an addition of IL-4. The clonogenicity of the nonadherent cells was not significantly influenced by IL-4. By addition of the media conditioned by OCI-AML1a cells in the presence of IL-4, the clonogenic cells growth of OCIAML1a cells was suppressed and adherent cells were markedly increased. The suppressive and differentiative effects on OCI/AML1a cells of the conditioned media and IL-4 itself were almost completely abolished by anti-IL-4 antibody. Furthermore, the neutralizing antibodies against transforming growth factor-beta 2 (TGF-beta 2), tumor necrosis factor-alpha (TNF-alpha), or IL-6 did not influence the effect of recombinant IL-4. Taken together, IL-4 was shown to suppress the growth and induce differentiation toward adherent macrophage-like cells of the G-CSF-dependent myeloblastic cell line. The effect of IL-4 may be direct, and not secondary via inducing production of other cytokines such as TGF-beta, TNF-alpha, or IL-6 by leukemic cells.

scholarly journals Human spleen cell generation of factors stimulating human pluripotent stem cell, erythroid, and myeloid progenitor cell growth

Blood ◽  
1985 ◽  
Vol 65 (4) ◽  
pp. 990-996
Author(s):  
I Fabian ◽  
D Douer ◽  
L Levitt ◽  
Y Kletter ◽  
PL Greenberg
Keyword(s):  
Stem Cell ◽  
Growth Factors ◽  
Spleen Cell ◽  
Pluripotent Stem Cell ◽  
Progenitor Cell ◽  
Target Cells ◽  
Nonspecific Esterase ◽  
Spleen Cells ◽  
Human Spleen ◽  
Nonadherent Cells

Mitogen-stimulated murine spleen cells produce humoral substances capable of supporting murine hematopoiesis and pluripotent stem cell proliferation in vitro. Thus, we evaluated conditioned media generated by human spleen cells (SCM) in the presence or absence of mitogens for factors stimulatory for human pluripotent (CFU-GEMM), erythroid (BFU- E), and myeloid (CFU-GM) precursors. Two and one half percent to 10% SCM stimulated proliferation of all three types of precursor cells from nonadherent buoyant human marrow target cells. Mitogen-stimulated SCM augmented CFU-GM (175% to 225%), whereas CFU-GEMM and BFU-E growth was essentially unchanged. Cell separation procedures used to determine which cells provided these microenvironmental stimuli indicated that nonadherent mononuclear spleen cells provided the bulk of the CSF-GM, whereas adherent cells (95% nonspecific esterase + monocyte- macrophages) and nonadherent cells provided similar proportions of CSF- mix and erythroid burst-promoting activity (BPA). The nonadherent cells generating high levels of CSF-mix, BPA, and CSF-GM were predominantly Leu-1-negative, ie, non-T, cells. In the presence or absence of mitogens, SCM was a more potent source (1.3- to 3.8-fold) than peripheral leukocyte CM of the growth factors for the three progenitor cell types. Specific in situ cytochemical stains for analyzing morphology of myeloid colonies demonstrated that SCM stimulated the proliferation of the same types and proportions of colonies as human placental CM, suggesting that these CMs may contain similar CSF-GMs. These data show the contribution of spleen cell subsets to the generation of hematopoietic growth factors and the responsiveness of these cells to various mitogenic stimuli.

Effects of 200cH medications on mice bone marrow cells and macrophages

2021 ◽  
Vol 10 (35) ◽  
pp. 75-76
Author(s):  
Carolina De Oliveira ◽  
Ana Paula R. Abud ◽  
Eneida Da Lozzo ◽  
Raffaello Di Bernardi ◽  
Simone De Oliveira ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Immune Cells ◽  
Bone Marrow Cells ◽  
Ex Vivo ◽  
Hematopoietic Lineage ◽  
Lineage Markers ◽  
Lachesis Muta ◽  
Nonadherent Cells ◽  
Marrow Cells

Paracelsus once wrote: "All things are poison and nothing is without poison, only the dose permits something not to be poisonous." Latter Hahnemann formulated the law of similars, preparations which cause certain symptoms in healthy individuals if given in diluted form to patients exhibiting similar symptoms will cure it. Highly diluted natural complexes prepared according to Hahnemann’s ancient techniques may represent a new form of immunomodulatory therapy. The lack of scientific research with highly diluted products led us to investigate the in vivo and in vitro actions of commonly used medications. Here we describe the results of experimental studies aimed at verifying the effects of Mercurius solubilis, Atropa Belladonna, Lachesis muta and Bryonia alba. All medications were at 200cH dilution. Animals were maintained for 7 days and were allowed to drink the medications, which were prepared in a way that the final dilution and agitation (200cH) was performed in drinking water. The medication bottle was changed and sucussed every afternoon. Co-culture of non treated mice bone marrow cells and in vitro treated peritoneal macrophages were also performed. After animal treatment the bone marrow cells were immunophenotyped with hematopoietic lineage markers on a flow cytometer. We have determined CD11b levels on bone marrow cells after culture and co-culture with treated macrophages and these macrophages were processed to scanning electron microscopy. We have observed by morphological changes that macrophages were activated after all treatments. Mercurius solubilis treated mice showed an increase in CD3 expression and in CD11b on nonadherent bone marrow cells after co-culture with in vitro treatment. Atropa Belladonna increased CD45R and decreased Ly-6G expression on bone marrow cells after animal treatment. Lachesis muta increased CD3, CD45R and, CD11c expression and decreased CD11b ex vivo and in nonadherent cells from co-culture. Bryonia alba increased Ly-6G, CD11c and CD11b expression ex vivo and when in co-culture CD11b was increased in adherent cells as well as decreased in nonadherent cells. With these results we have demonstrated that highly diluted medications act on immune cells activating macrophages, and changing the expression profile of hematopoietic lineage markers. Highly diluted medications are less toxic and cheaper than other commonly used medications and based on our observations, it is therefore conceivable that this medications which are able to act on bone marrow and immune cells may have a potential therapeutic use in clinical applications in diseases were the immune system is affected and also as regenerative medicine as it may allow proliferation and differentiation of progenitor cells.

scholarly journals Isolation, Maintenance and Differentiation of Primary Tracheal Basal Cells from Adult Rhesus Macaque

2019 ◽  
Vol 2 (4) ◽  
pp. 79
Author(s):  
Anna E. Engler ◽  
Gustavo Mostoslavsky ◽  
Lisa Miller ◽  
Jason R. Rock
Keyword(s):  
Rhesus Macaque ◽  
Rhesus Macaques ◽  
Enzymatic Digestion ◽  
Basal Cells ◽  
Early Passage ◽  
Long Term Storage ◽  
Air Liquid Interface ◽  
Nonadherent Cells ◽  
Level 2

In this report, we describe methodologies for the isolation and culture of primary rhesus macaque tracheal basal cells, their cryopreservation, long term storage and differentiation. These are comparable to state-of-the-art protocols that have been developed for mouse and human airway basal cells. This method is based on the use of proprietary media, providing an easily reproducible and applicable protocol for usage in biosafety level 2 (BSL2) settings. Tracheas from rhesus macaques were isolated after animal euthanasia and subjected to enzymatic digestion overnight. Cells of the epithelial layer were scraped off of the trachea for cell culture. Twenty-four hours after plating basal cells had attached and nonadherent cells were removed. First passages of basal cells can be frozen for early passage storage in liquid nitrogen or propagated and differentiated on an air–liquid interface and in a tracheosphere assay up to passage seven. This protocol provides a platform for the analysis of basal cells from a close evolutionary relative to humans.

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