scholarly journals Enhancement of In Vitro Growth of Pathogenic Bacteria by Norepinephrine: Importance of Inoculum Density and Role of Transferrin

2006 ◽  
Vol 72 (7) ◽  
pp. 5097-5099 ◽  
Author(s):  
Phyllis M. O'Donnell ◽  
Hernan Aviles ◽  
Mark Lyte ◽  
Gerald Sonnenfeld
Keyword(s):  
Bacterial Growth ◽  
Pathogenic Bacteria ◽  
Inoculum Density ◽  
In Vitro Growth ◽  
Stress Hormone ◽  
Serum Free ◽  
Free Media

ABSTRACT Norepinephrine is a stress hormone that enhances bacterial growth. We examined the effects of a small inoculum on the norepinephrine-induced growth of species previously reported to be unaffected by norepinephrine. The results indicated that a reduced inoculum density is essential for observing norepinephrine-induced effects. Additional studies using serum-free media suggested that transferrin plays a role in norepinephrine-induced growth.

268 ROLE OF ATPase MOTOR CYTOPLASMIC DYNEIN AND PRIMARY CILIA IN TESTICULAR MORPHOGENESIS

2015 ◽  
Vol 27 (1) ◽  
pp. 223
Author(s):  
C. Dores ◽  
I. Dobrinski
Keyword(s):  
Primary Cilia ◽  
Somatic Cells ◽  
Treated Group ◽  
Cytoplasmic Dynein ◽  
Control Group ◽  
Tubule Formation ◽  
Serum Free ◽  
Free Media

In vertebrates, the primary cilium is a nearly ubiquitous organelle present in somatic cells, but little is known about its function in the male gonad. We investigated the role of primary cilia in testis cells using in vitro formation of seminiferous tubules and in vitro culture of testicular somatic cells by inhibiting the primary cilium with CiliobrevinD, a cell-permeable, reversible chemical modulator that inhibits the major component of the organelle: ATPase motor cytoplasmic dynein. We analysed in vitro cultures for the presence of primary cilia and the activation of hedgehog signalling through translocation of Gli2 to the nuclei; in vitro tubule formation was evaluated by length and width of tubules formed. Methods: testicular cells were harvested from neonatal pigs by 2-step enzymatic digestion. Cells (50 × 106 mL–1) were plated on 100 mm Petri dishes in 15 mL of DMEM + 5% FBS + 50 U of penicillin and incubated at 37°C in 5% CO2 in air overnight, cells remaining in suspension and those slightly attached were removed and the somatic cells attached were trypsinized to obtain a single cell suspension, and then submitted to two different protocols: in vitro culture (A) or in vitro tubule formation (B), n = 5 replicates each. For A, somatic cells were replated on coverslips in 24-well plates and cultured in serum free media for 48 h, then for the treated group, 10 mM of CiliobrevinD was added for 24 h, attached cells from control and treated groups were fixed in 4% PFA and characterised by immunocytochemistry for ARL13B, Vimentin, and Gli2. For B: 1 × 106 cells were added to 24-well plates coated with 1 : 1 diluted Matrigel, the control group was kept in serum free media and to the treated group was added 20 mM CiliobrevinD at Day 0. Results: A) primary cilia were present in 89.3 ± 2.3% of cells cultured in serum-free media for the control group and Gli2 was located in the nuclei of 90.2 ± 1.2% of cells; in the CiliobrevinD-treated group the percentage of primary cilia decreased (P < 0.05) to 3.1 ± 2.5% and nuclear Gli2 to 3.9 ± 0.7; B) tubules formed in the control group were significantly longer and wider than the ones formed when CiliobrevinD was added (9.91 ± 0.35 v. 5.540 ± 1.08 mm and 339.8 ± 55.78 v. 127.2 ± 11.9 µm, respectively, P < 0.05 by Student's t-test). In conclusion, the inhibition of ATPase motor cytoplasmic dynein perturbs formation of primary cilia in testicular somatic cells, blocks Hedgehog signalling, and impairs in vitro tubule formation. Therefore, primary cilia on testicular somatic cells appear to be essential for testicular morphogenesis.Research was supported by 5 R01 OD016575-13.

THE ROLE OF CYTOKININS AND ETHYLENE INHIBITORS ON LENTICEL HYPERTROPHY GENERATION AND ETHYLENE PRODUCTION IN IN VITRO CULTURES OF POPULUS EUPHRATICA OLIVIER

1995 ◽  
Vol 43 (4) ◽  
pp. 339-345 ◽  
Author(s):  
M.D. Lledó ◽  
M.B. Crespo ◽  
J.B. Amo-Marco
Keyword(s):  
Ethylene Production ◽  
Populus Euphratica ◽  
In Vitro Cultures ◽  
Nodal Segments ◽  
Ethylene Inhibitors ◽  
Se Spain ◽  
Ethylene Formation ◽  
Free Media

Populus euphratica Olivier is native to the Irano—Turanian areas (Middle East). Elche (Alicante province, SE Spain) is known to be its only European location. Nodal segments from root shoots were established in vitro in a Murashige and Skoog medium supplemented with several cytokinins. Ethylene inhibitors AgNO3 and CoCl2 were used in combination with kinetin. Hormone-free media supplemented with sucrose (20–60 mg 1−1) was also tested. Ethylene was measured by gas chromatography, and both the percentage of sprouting shoots and lenticel hypertrophy in cultures were recorded. Ethylene production was higher in cultures supplemented with cytokinins (especially with meta-topolin), with high sprouting percentages, and lenticel hypertrophy. In cultures supplemented with 6-benzylaminopurine or 6-(γ,γ,-dimethylallylamino)-purine, ethylene production was lower and explants looked unhealthy. Ethylene formation was inhibited in cultures supplemented with AgNO3 (1 mg 1−1), which also decreased percentage of sprouting buds and lenticel hypertrophy.

TMOD-21. A NOVEL IN-VITRO METHOD TO MODEL MACROPHAGES IN GLIOBLASTOMA

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi220-vi220
Author(s):  
Hasan Alrefai ◽  
Andee Beierle ◽  
Lauren Nassour ◽  
Nicholas Eustace ◽  
Zeel Patel ◽  
...  
Keyword(s):  
Cell Types ◽  
In Vitro Models ◽  
Tumor Initiating Cells ◽  
Serum Free ◽  
Brain Tumor Initiating Cells ◽  
Multiple Cell ◽  
Anti Inflammatory ◽  
Free Media ◽  
Inflammatory Macrophages

Abstract BACKGROUND The GBM tumor microenvironment (TME) is comprised of a plethora of cancerous and non-cancerous cells that contribute to GBM growth, invasion, and chemoresistance. In-vitro models of GBM typically fail to incorporate multiple cell types. Others have addressed this problem by employing 3D bioprinting to incorporate astrocytes and macrophages in an extracellular matrix; however, they used serum-containing media and classically polarized anti-inflammatory macrophages. Serum has been shown to cause GBM brain-tumor initiating cells to lose their stem-like properties, highlighting the importance of excluding it from these models. Additionally, tumor-associated macrophages (TAMs) do not adhere to the traditional M2 phenotype. METHODS THP-1 monocytes and normal human astrocytes (NHAs) were transitioned into serum-free HL-1 and neurobasal-based media, respectively. Monocytes were stimulated towards a macrophage-like state with PMA and polarized by co-culturing them with GBM patient-derived xenograft(PDX) lines, using a transwell insert. CD206 expression was used to validate polarization and a cytokine array was used to characterize the cells. RESULTS There was no difference in proliferation rates at 72 hours for THP-1 monocytes grown in serum-free HL-1 media compared to serum-containing RPMI 1640 (p &gt; 0.95). Macrophages polarized via transwell inserts expressed the lymphocyte chemoattractant protein, CCL2, whereas resting(M0), pro-inflammatory(M1), and anti-inflammatory(M2) macrophages did not. Additionally, these macrophages expressed more CXCL1 and IL-1ß relative to M1 macrophages. We have also demonstrated a method to maintain a tri-culture model of GBM PDX cells, NHAs, and TAMs in a serum-free media that supports the growth/maintenance of all cell types. CONCLUSIONS We have demonstrated a novel method by which we can polarize macrophages towards a tumor-supportive phenotype that differs in cytokine expression from traditionally polarized macrophages. This higher-fidelity method of modeling TAMs in GBM can aid in the development of targeted therapeutics that may one day enter the clinic in hopes of improving outcomes in GBM.

Role of Organic Phosphate in Mineralization of Bone in vitro

1981 ◽  
Vol 60 (C) ◽  
pp. 1586-1586 ◽  
Author(s):  
H. C. Tenenbaum
Keyword(s):  
Defined Medium ◽  
Organic Phosphate ◽  
Limiting Factor ◽  
Serum Free ◽  
Osteogenic Cells ◽  
Inorganic Phosphates

Calvarial periostea were dissected from 17-day-old embryonic chicks and folded with the osteogenic cells in apposition. The folded explains were cultured for up to six d on serum and plasma clots or in serum-free hormone-supplemented completely-defined medium. Osteoid consistently formed in such cultures in both types of media, and this osteoid mineralized when appropriate levels of β-glycerophosphate were added to each type of medium. The data presented suggest that the levels of organic phosphate might be more important than inorganic phosphates as a limiting factor in the initiation of mineralization of bone in vitro.

Catecholamines and in vitro growth of pathogenic bacteria: enhancement of growth varies greatly among bacterial species

Life Sciences ◽  
2003 ◽  
Vol 73 (12) ◽  
pp. 1527-1535 ◽  
Author(s):  
Tesfaye Belay ◽  
Hernan Aviles ◽  
Monique Vance ◽  
Kimberly Fountain ◽  
Gerald Sonnenfeld
Keyword(s):  
Pathogenic Bacteria ◽  
Bacterial Species ◽  
In Vitro Growth

Role of kallikrein-kininogen system in insulin-stimulated glucose transport after muscle contractions

2002 ◽  
Vol 92 (2) ◽  
pp. 657-664 ◽  
Author(s):  
C. L. Dumke ◽  
J. Kim ◽  
E. B. Arias ◽  
G. D. Cartee
Keyword(s):  
Glucose Transport ◽  
Serum Proteins ◽  
Muscle Contractions ◽  
Rat Serum ◽  
Serum Free ◽  
Bradykinin Receptor ◽  
Contraction Effect ◽  
Kallikrein Inhibitors

Serum proteins [molecular weight (MW) > 10,000] are essential for increased insulin-stimulated glucose transport after in vitro muscle contractions. We investigated the role of the kallikrein-kininogen system, including bradykinin, which is derived from kallikrein (MW > 10,000)-catalyzed degradation of serum protein kininogen (MW > 10,000), on this contraction effect. In vitro electrical stimulation of rat epitrochlearis muscles was performed in 1) rat serum ± kallikrein inhibitors; 2) human plasma (normal or kallikrein-deficient); 3) rat serum ± bradykinin receptor-2 inhibitors; or 4) serum-free buffer ± bradykinin. 3- O-methylglucose transport (3-MGT) was measured 3.5 h later. Serum ± kallikrein inhibitors tended ( P = 0.08) to diminish postcontraction insulin-stimulated 3-MGT. Contractions in normal plasma enhanced insulin-stimulated 3-MGT vs. controls, but contractions in kallikrein-deficient plasma did not. Supplementing rat serum with bradykinin receptor antagonist HOE-140 during contraction did not alter insulin-stimulated 3-MGT. Muscles stimulated to contract in serum-free buffer plus bradykinin did not have enhanced insulin-stimulated 3-MGT. Bradykinin was insufficient for postcontraction-enhanced insulin sensitivity. However, results with kallikrein inhibitors and kallikrein-deficient plasma suggest kallikrein plays a role in this improved insulin action.

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