The mitogenic activity of lipopolysaccharide for spleen cells from germfree, conventional, and gnotobiotic rats

1979 ◽  
Vol 25 (9) ◽  
pp. 1087-1093 ◽  
Author(s):  
Carol Wells ◽  
Edward Balish
Keyword(s):  
Spleen Cell ◽  
Lipid A ◽  
Mitogenic Activity ◽  
Adherent Cells ◽  
Spleen Cells ◽  
E Coli ◽  
Blastogenic Response ◽  
Pseudomonas Species ◽  
Nonadherent Cells ◽  
Germfree Rats

Spleen cells from germfree rats, conventionally reared rats, and gnotobiotic rats associated with two Pseudomonas species gave no positive blastogenic response when incubated with each of four lipopolysaccharide (LPS) preparations from Escherichia coli, with glycolipid extracted from Salmonella minnesota R595 or with S. minnesota R595 lipid A. However, spleen cell preparations from athymic mice demonstrated a positive blastogenic response when incubated with E. coli LPS. Removal of adherent cells from germfree and conventional-flora rat spleen cells did not increase the mitogenic activity of LPS for nonadherent cells (< 0.5% esterase-positive cells). All rat spleen cell preparations gave positive blastogenic responses to phytohemagglutinin and concanavalin A. This study indicates that LPS may not be a mitogenic agent for rat spleen 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.

Dual effect of meningococcal antigens on a T cell dependent immune response

1983 ◽  
Vol 29 (12) ◽  
pp. 1619-1625 ◽  
Author(s):  
Brian G. Sparkes
Keyword(s):  
Immune Response ◽  
Inhibitory Activity ◽  
Sheep Erythrocyte ◽  
Subsequent Development ◽  
Plaque Formation ◽  
Adherent Cells ◽  
Spleen Cells ◽  
Dual Effect ◽  
Adjuvant Activity ◽  
Nonadherent Cells

Meningococcal antigens (MA) showed adjuvant activity when administered to mice at the same time as antigen (sheep erythrocyte (SE)), by increasing the splenocyte plaque-forming response in a dose-related manner. However, when SE were given 1 day after MA administration, the subsequent plaque formation was diminished from normal in proportion to the dose of MA injected. Splenocytes taken from mice up to 5 days after MA injection actively inhibited plaque formation when mixed with splenocytes immunized with SE 4 days earlier. Two days after MA injection the nonspecific inhibition of plaque formation was mainly due to adherent spleen cells, while at 5 days nonadherent cells had acquired the inhibitory activity. It appears that it is the degree of activation of adherent cells resulting from the timing and dosage of MA which modulates the subsequent development and secretion of antibody-forming cells.

Soluble factors from rabbit spleen cells kill and lyse Treponema pallidum in vitro

1990 ◽  
Vol 36 (10) ◽  
pp. 711-717 ◽  
Author(s):  
Thomas J. Fitzgerald ◽  
Barbara J. Elmquist
Keyword(s):  
Cytochalasin B ◽  
Treponema Pallidum ◽  
Adherent Cells ◽  
Spleen Cells ◽  
Soluble Factors ◽  
Macrophage Phagocytosis ◽  
Immune Mediated ◽  
Nonadherent Cells ◽  
Immune Rabbit

Antibody and complement immobilize (kill) Treponema pallidum in vitro. Recent evidence also documents immobilization by soluble factors released by activated macrophages and lymphocytes. Immune-mediated lysis of treponemes, however, has not been reported. The findings in this paper focus on apparent treponemal lysis by rabbit splenic cell preparations. Using cells from animals infected testicularly for 9 to 12 days, unfractionated splenic preparations, as well as adherent and nonadherent preparations, killed and lysed T. pallidum. Phagocytosis alone could not explain the detrimental effects of adherent cells. When cytochalasin B was used to block phagocytosis, decreases in treponemal numbers were still detected. In related studies, immune rabbit sera did not enhance treponemicidal activity of the adherent cells. To assess the specificity of these reactions, T. pallidum was incubated with two monocyte-like cell lines (human U937 and mouse P388D1). Neither cell line was detrimental, and treponemal numbers were not lowered. The soluble nature of the treponemicidal factors from adherent and nonadherent preparations was shown by physically separating these cells from the organisms and demonstrating treponemal killing and lysis. In summary, clearance of T. pallidum from infected tissues is probably at least partially attributed to macrophage phagocytosis. Our findings suggest another mechanism involving lytic factors secreted by activated adherent and nonadherent cells. Key words: syphilis, splenic treponemicidal activity, Treponema pallidum.

scholarly journals POTENTIATION OF THE T-LYMPHOCYTE RESPONSE TO MITOGENS

1972 ◽  
Vol 136 (1) ◽  
pp. 143-155 ◽  
Author(s):  
Igal Gery ◽  
Byron H. Waksman
Keyword(s):  
T Cells ◽  
Bone Marrow Cells ◽  
Mouse Spleen ◽  
Adherent Cells ◽  
Spleen Cells ◽  
Con A ◽  
Cell Responses ◽  
Donor Cells ◽  
Peripheral Leukocytes ◽  
Nonadherent Cells

Effective supernatants (SUP), which potentiate mouse T-cell responses to phytohemagglutin (PHA), are obtained from cells of several species (human, rabbit, rat, mouse) and indeed from syngeneic spleen, thymus, or bone marrow cells. Unstimulated cells release some SUP activity but more is produced after stimulation. Lipopolysaccharide (LPS) produced very active SUP in all cultures tested. PHA was similarly active on human leukocytes only, whereas concanavalin A (Con A) gave highly efficient SUP only with mouse spleen cells. SUP production is not correlated with a mitotic response of the donor cells and is observed in cultures unable to respond mitotically to the stimulant. Adherent mouse spleen cell populations, consisting largely or entirely of macrophages, produce active SUP, while nonadherent cells do not. Similarly, purification of human peripheral leukocytes on nylon columns, with removal of macrophages and other adherent cells, destroys their ability to produce SUP. The importance of indirect effects in stimulating mitotic responses of T cells is emphasized by the fact that the mitotic response of mouse thymocytes to LPS and its ability to potentiate the response of these cells to PHA disappears with removal of adherent cells from the thymocyte population. Conversely the production of SUP from spleen cells stimulated by Con A requires the presence of T cells.

scholarly journals Isolation of a lipid a bound polypeptide responsible for LPS- initiated mitogenesis of C3H/HeJ spleen cells

1976 ◽  
Vol 144 (3) ◽  
pp. 840-846 ◽  
Author(s):  
DC Morrison ◽  
SJ Betz ◽  
DM Jacobs
Keyword(s):  
Lipid A ◽  
Critical Role ◽  
Gel Filtration ◽  
Polymyxin B ◽  
Mitogenic Activity ◽  
Partial Purification ◽  
Spleen Cells ◽  
Phenol Treatment ◽  
Bacterial Lipopolysaccharides

The experiments by Sultzer and Nilsson (1), and later by Watson and Riblet (2), established that spleen cells from the C3H/HeJ strain of mouse were refractory to the mitogenic effects of bacterial lipopolysaccharides (LPS). More recently, however, experiments from our laboratory (3) demonstrated that spleen cells from C3H/HeJ mice were in fact responsive to some preparations of LPS but not to others, and that the method of extraction played a critical role in determining activity. In particular, preparations of LPS prepared by extraction with aqueous butanol had potent mitogenic activity. Our data showed that the mitogenic activity of such positive preparations of LPS coisolated with the LPS during gel filtration chromatography and subsequent equilibrium banding on CsCl. In addition, lipid A isolated from positive preparations of LPS was also capable of stimulating C3H/HeJ spleen cells. Taken together, these experiments provided rather convincing data that it was the LPS (in particular the lipid A) itself, or some contaminant very tightly bound to the lipid A, which was responsible for its biological activity. We further demonstrated that treatment of positive preparations of LPS with hot phenol rendered such preparations nonmitogenic for C3H/HeJ spleens, yet activity for other strains was only moderately decreased. These experiments would suggest either that the phenol treatment chemically alters the lipid A region of the LPS molecule or that such treatment removes the putative tightly bound contaminant responsible for C3H/HeJ mitogenesis. In the experiments reported here, we have explored in greater detail the role of lipid A in the stimulation of C3H/HeJ spleen cells. For these experiments we have utilized our earlier observations that the antibiotic polymyxin B forms a highly stable molecular complex with the lipid A region of LPS (4), and that such polymyxin B-LPS complexes are unable to mitogenically stimulate B lymphocytes (5). In addition, we have attempted to distinguish between the two potential modes of action of phenol on LPS, namely, the chemical alteration of the lipid A or the removal of a tightly bound contaminant by phenol treatment. The results of the experiments we report here support the interpretation that mitogenic activity of positive preparations of LPS is associated with a low mol wt phenol soluble polypeptide of approximately 10,000 mol wt. After partial purification, this polypeptide intitiates a significant mitogenic response at concentrations as low as 10 μg/ml. We conclude that the C3H/HeJ strain of mouse is a true nonresponder to the stimulatory effects of the lipid A region of LPS.

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

Abstract 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.

scholarly journals Regulation of the antibody response to type III pneumococcal polysaccharide by contrasuppressor T cells.

1984 ◽  
Vol 160 (1) ◽  
pp. 42-54 ◽  
Author(s):  
H Braley-Mullen
Keyword(s):  
T Cells ◽  
Spleen Cell ◽  
Adherent Cells ◽  
Spleen Cells ◽  
Suppressor T Cells ◽  
Membrane Component ◽  
Vicia Villosa ◽  
Type Iii ◽  
Cell Fractions ◽  
Specific Suppressor T Cells

A soluble membrane component of type III pneumococcal polysaccharide-coupled spleen cells (S3-SCSM) induces S3-specific suppressor T cells (Ts) in mice. These Ts can be detected only if mice are pretreated with cyclophosphamide (Cy) or if cells adherent to the lectin Vicia villosa are removed from the spleen cell population prior to transfer. The V. villosa-adherent spleen cells from mice injected with S3-SCSM could abrogate suppression mediated by Ts induced by S3-SCSM in Cy-treated mice. The V. villosa-adherent contrasuppressor cells were shown to be T cells that were I-J+ and of the Lyt-1 phenotype. Contrasuppressor T cells (Tcs) were not present in V. villosa-adherent spleen cell fractions obtained from normal mice, from mice injected with polyvinylpyrrolidone-coupled spleen cells, or from Cy-treated mice injected with S3-SCSM, i.e., mice in which Ts activity is dominant. The V. villosa-adherent cells that abrogated the activity of Ts induced by S3-SCSM in Cy-treated mice did not abrogate suppression mediated by a different subset of S3-specific Ts, suggesting that the Tcs described here do not have activity against all Ts subsets. The ability of S3-SCSM to activate Tcs in normal mice provides an explanation for the inability to detect S3-specific Ts in several previous studies.

scholarly journals Granulibacter bethesdensis, a Pathogen from Patients with Chronic Granulomatous Disease, Produces a Penta-Acylated Hypostimulatory Glycero-d-talo-oct-2-ulosonic Acid–Lipid A Glycolipid (Ko-Lipid A)

2021 ◽  
Vol 22 (7) ◽  
pp. 3303
Author(s):  
Artur Muszyński ◽  
Kol A. Zarember ◽  
Christian Heiss ◽  
Joseph Shiloach ◽  
Lars J. Berg ◽  
...  
Keyword(s):  
Chronic Granulomatous Disease ◽  
Human Blood ◽  
Lipid A ◽  
Acid Resistance ◽  
Strong Acid ◽  
Granulomatous Disease ◽  
E Coli ◽  
Phagocyte Nadph Oxidase ◽  
Acyl Chains ◽  
Ulosonic Acid

Granulibacter bethesdensis can infect patients with chronic granulomatous disease, an immunodeficiency caused by reduced phagocyte NADPH oxidase function. Intact G. bethesdensis (Gb) is hypostimulatory compared to Escherichia coli, i.e., cytokine production in human blood requires 10–100 times more G. bethesdensis CFU/mL than E. coli. To better understand the pathogenicity of G. bethesdensis, we isolated its lipopolysaccharide (GbLPS) and characterized its lipid A. Unlike with typical Enterobacteriaceae, the release of presumptive Gb lipid A from its LPS required a strong acid. NMR and mass spectrometry demonstrated that the carbohydrate portion of the isolated glycolipid consists of α-Manp-(1→4)-β-GlcpN3N-(1→6)-α-GlcpN-(1⇿1)-α-GlcpA tetra-saccharide substituted with five acyl chains: the amide-linked N-3′ 14:0(3-OH), N-2′ 16:0(3-O16:0), and N-2 18:0(3-OH) and the ester-linked O-3 14:0(3-OH) and 16:0. The identification of glycero-d-talo-oct-2-ulosonic acid (Ko) as the first constituent of the core region of the LPS that is covalently attached to GlcpN3N of the lipid backbone may account for the acid resistance of GbLPS. In addition, the presence of Ko and only five acyl chains may explain the >10-fold lower proinflammatory potency of GbKo–lipidA compared to E. coli lipid A, as measured by cytokine induction in human blood. These unusual structural properties of the G.bethesdensis Ko–lipid A glycolipid likely contribute to immune evasion during pathogenesis and resistance to antimicrobial peptides.

scholarly journals Immunologic properties of bacterial lipopolysaccharide (LPS). II. The unresponsiveness of C3H/HeJ Mouse spleen cells to LPS-induced mitogenesis is dependent on the method used to extract LPS.

1975 ◽  
Vol 142 (6) ◽  
pp. 1488-1508 ◽  
Author(s):  
B J Skidmore ◽  
D C Morrison ◽  
J M Chiller ◽  
W O Weigle
Keyword(s):  
B Cells ◽  
Mouse Strain ◽  
Structural Integrity ◽  
Mitogenic Activity ◽  
Bacterial Lipopolysaccharide ◽  
Mouse Strains ◽  
Striking Difference ◽  
Spleen Cells ◽  
Destructive Effect

The C3H/HeJ mouse strain, previously shown to be a nonresponder to bacterial lipopolysaccharide (LPS)-induced mitogenesis in vitro, was demonstrated by the present studies to be competent to respond mitogenically to LPS, but only to LPS preparations obtained by selected extraction methods. These preparations appear to be confined to LPS isolated by mild extraction techniques, such as TCA or butanol. In contrast, those obtained by techniques utilizing phenol were only weakly stimulatory or completely nonstimulatory for spleen cells from the C3H/HeJ. All LPS preparations tested, on the other hand, were highly stimulatory for cells from another mouse strain, namely the C3H/St. The critical importance of the method of extraction of LPS on its mitogenic activity for C3H/HeJ cells was stressed by experiments in which LPS was prepared from Escherichia coli K235 using either of two procedures. In these experiments, phenol-extracted LPS, although mitogenic in the C3H/St, was completely nonstimulatory in the C3H/HeJ; whereas, butanol-extracted LPS was highly stimulatory in both strains of mice. This striking difference was attributed to a destructive effect of phenol on LPS, as demonstrated by the fact that treatment of butanol LPS with phenol resulted in a total loss of its mitogenic activity in the C3H/HeJ, but in only a partial loss in the C3H/St. In general, the mitogenic response observed with selected LPS preparations in the C3H/HeJ was quantitatively lower and more transient than that seen with the C3H/St, although qualitatively these responses appeared to be similar. This was evidenced by the observation that in both mouse strains LPS was a specific mitogen for B cells, a property which was also attributed in both strains to the same distinct structural region of the LPS molecule, that is lipid A. A preparation of LPS that failed to stimulate B cells from the C3H/HeJ nonetheless had the capacity to block activation of these B cells by a stimulatory preparation of LPS. These results strongly suggest that mitogenic stimulation of B cells by LPS is a function of the structural integrity of both the LPS molecule and putative B-cell receptors for LPS.

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.

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