scholarly journals Direct inhibitory effect of caffeine on viability, synthesis activity and gene expression in cultures of chondrocytes extracted from the articular cartilage of rats

2019 ◽  
Vol 71 (2) ◽  
pp. 509-520
Author(s):  
A.M.S. Reis ◽  
K.P. Oliveira ◽  
I.H.F. Paula ◽  
A.P. Silva ◽  
J.F. Tarragô ◽  
...  
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Articular Cartilage ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Collagen Synthesis ◽  
Inhibitory Effect ◽  
Direct Inhibitory Effect ◽  
Synthesis Activity ◽  
Safranin O

ABSTRACT The aim of this study was to evaluate the effect of concentrations of caffeine on the viability, synthesis activity and gene expression in cultures of chondrocytes. Extracted articular cartilage from the femurs and tibias of 15 Wistar rats at three days old to isolate chondrocytes. Chondrocytes were cultured in chondrogenic medium (control) or supplemented with caffeine (0.5, 1.0, 2.0mM). Cell viability, alkaline phosphatase activity and collagen synthesis were assessed using colorimetric assays at 7, 14, 21 days. The chondrocyte cultures of all groups grown under coverslips were stained with hematoxylin-eosin to determine the percentage of cells/field and with PAS, safranin O, alcian blue to determine the percentage of matrix chondrogenic/field at 21 days. The expressions of gene transcripts for aggrecan, collagen-II, Sox-9, Runx-2 and alkaline phosphatase were also evaluated by RT-PCR at 21 days. The means were compared using Student-Newman-Keuls. Caffeine significantly reduced the conversion of MTT to formazan, percentage of cells/field, collagen synthesis, alkaline phosphatase activity, synthesis of PAS+, safranin O+ and alcian blue+ chondrogenic matrix, and the expression of aggrecan, Sox-9 and II collagen. It is concluded that caffeine at concentrations of 0.5, 1.0, 2.0mM has a direct inhibitory effect on chondrogenesis in cultures of chondrocytes from rats.

Effect of Alkaline and Acid Phosphatases on Clotting:

1966 ◽  
Vol 15 (03/04) ◽  
pp. 365-380 ◽  
Author(s):  
P. G Iatridis ◽  
J. H Ferguson
Keyword(s):  
Alkaline Phosphatase ◽  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Inhibitory Effect ◽  
Factor Ix ◽  
Acid Phosphatases ◽  
Acid And Alkaline Phosphatase ◽  
Direct Inhibitory Effect ◽  
Alkaline And Acid Phosphatases

SummaryAlkaline phosphatase is found to enhance the activation of factor IX by SF. The correlation of the distribution of alkaline phosphatase in electrophoretic fractions with the clotting tests suggest that the “beta” fraction contains the responsible factor for the acceleratory effect of alkaline phosphatase on clotting.Acid phosphatase, while not exerting a direct inhibitory effect on SF, does enhance the plasmatic anti-SF activity. The “beta” and “F-gamma” fractions seem to contain the responsible factor of acid phosphatase for the plasmatic anti-SF enhancement.SF preparation has no acid or alkaline phosphatase activity.A tentative schema is proposed to explain the effects of acid and alkaline phosphatase on clotting.

The influence of titanium surfaces treated by alkalis on macrophage and osteoblast-like cell adhesion and gene expression in vitro

RSC Advances ◽  
2015 ◽  
Vol 5 (99) ◽  
pp. 81378-81387 ◽  
Author(s):  
Ting Ma ◽  
Xi-Yuan Ge ◽  
Sheng-Nan Jia ◽  
Xi Jiang ◽  
Yu Zhang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Cell Adhesion ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Related Gene ◽  
Titanium Surfaces

The effect of alkali-treated titanium surfaces on inflammation-related gene expression of macrophages and alkaline phosphatase activity of osteoblast-like cells.

Acidosis inhibits osteoblastic and stimulates osteoclastic activity in vitro

1992 ◽  
Vol 262 (3) ◽  
pp. F442-F448 ◽  
Author(s):  
N. S. Krieger ◽  
N. E. Sessler ◽  
D. A. Bushinsky
Keyword(s):  
Alkaline Phosphatase ◽  
Metabolic Acidosis ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Collagen Synthesis ◽  
Calcium Flux ◽  
Osteoclastic Activity ◽  
Glucuronidase Activity

Metabolic acidosis induces net calcium flux (JCa) from cultured neonatal mouse calvariae through physicochemical and cell-mediated mechanisms. To determine the role of osteoblasts in acid-induced JCa, collagen synthesis and alkaline phosphatase activity were assessed in calvariae incubated in reduced pH and bicarbonate medium, a model of metabolic acidosis (Met), and compared with controls (Ctl). Collagen synthesis fell from 30.5 +/- 1.1 in Ctl to 25.1 +/- 0.4% with Met, and alkaline phosphatase decreased from 403 +/- 25 in Ctl to 298 +/- 21 nmol Pi.min-1.mg protein-1 with Met. During acidosis JCa was correlated inversely with percent collagen synthesis (r = -0.743, n = 11, P = 0.009) and with alkaline phosphatase activity (r = -0.453, n = 22, P = 0.034). To determine the role of osteoclasts in acid-induced JCa, osteoclastic beta-glucuronidase activity was determined in Ctl and Met in the absence or presence of the osteoclastic inhibitor calcitonin (CT, 3 x 10(-9) M). Met increased beta-glucuronidase (5.9 +/- 0.2) compared with Ctl (4.6 +/- 0.3 micrograms phenolphthalein released.bone-1.h-1), whereas CT inhibited beta-glucuronidase in both Ctl and Met (3.1 +/- 0.2 and 3.5 +/- 0.3, respectively). During acidosis JCa was correlated directly with beta-glucuronidase activity (r = 0.683, n = 42, P less than 0.001). Thus the cell-mediated component of JCa during acidosis in vitro appears to result from a combination of inhibited osteoblastic and stimulated osteoclastic activity.

OSTEOPLASTIC PROPERTIES OF MULTIPOTENT MESENCHYMAL STROMAL CELLS OF ADIPOSE TISSUE

2021 ◽  
Vol 74 (10) ◽  
pp. 2374-2378
Author(s):  
Andriy Bambuliak ◽  
Nataliia Kuzniak ◽  
Valentyna Honcharenko ◽  
Marianna Ostafiychuk ◽  
Alina Palamar
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Adipose Tissue ◽  
Phosphatase Activity ◽  
Mesenchymal Stromal Cells ◽  
Alkaline Phosphatase Activity ◽  
Stromal Cells ◽  
Multipotent Mesenchymal Stromal Cells ◽  
Tissue Samples ◽  
Medical University

The aim: Determining the ability of samples based on MMSC – AT differentiating in the osteogenic direction. Materials and methods: The study was conducting at Bukovinian State Medical University, Chernivtsi, Ukraine. Adipose tissue samples were obtaining from the neck of 60 experimental animals (white Wistar rats). Multipotent mesenchymal stromal cells of adipose tissue were obtained by grinding adipose tissue of rats in 0.1% collagenase 1A . Alkaline phosphatase activity was assessing by using the Alkaline Phosphatase Detection Kit (Sigma, USA) according to the manufacturer’s protocol. Osteopontin gene expression was determining by immunocytochemical method. To determine the mRNA used the PCR method, which is associated with reverse transcription (RT-PCR) in the area of quantification of gene expression to the marker BGP. Results: On the 21st day of observations, the expression of mRNA encoding the BGP gene decreased in samples № 1 and № 3 to 35,800 ± 420.0 copies and to 35,000 ± 400.0 copies, p1<0.01, p>0.05. Also was observing growth of copies of the BGP gene in samples № 2 and № 4 in 2.1, р<0.01 and 2.2 times, р-р2<0.05, relative to the data in sample № 1. Conclusions: Comparative study of osteoplastic properties samples MMSC-AT showed that a larger number of cells differentiate into the osteoblasts in samples containing MMSC-AT + PRP (№ 2) and MMSC-AT + PRP + «Kolapan» (№ 4). This has been proven higher alkaline phosphatase activity, higher levels osteopontin expression, and higher levels BGP gene expression.

Stimulated osteoclastic and suppressed osteoblastic activity in metabolic but not respiratory acidosis

1995 ◽  
Vol 268 (1) ◽  
pp. C80-C88 ◽  
Author(s):  
D. A. Bushinsky
Keyword(s):  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Collagen Synthesis ◽  
Cell Function ◽  
Respiratory Acidosis ◽  
Bone Cell ◽  
Calcium Efflux ◽  
Osteoblastic Activity ◽  
Glucuronidase Activity

When bone is cultured in acidic medium produced by a reduced bicarbonate concentration ([HCO(3-)]), a model of metabolic acidosis, there is greater net calcium efflux than when the same decrement in pH is produced by an increased partial pressure of carbon dioxide (PCO2), a model of respiratory acidosis. To determine the effects of metabolic and respiratory acidosis on bone cell function we cultured neonatal mouse calvariae for 48 h under control conditions (pH approximately 7.40, PCO2 approximately 41 mmHg, [HCO(3-)] approximately 25 meq/l) or under isohydric acidic conditions simulating metabolic (pH approximately 7.09, [HCO(3-)] approximately 12) or respiratory (pH approximately 7.10, PCO2 approximately 86) acidosis and measured osteoblastic collagen synthesis and alkaline phosphatase activity and osteoclastic beta-glucuronidase activity. Collagen synthesis was inhibited by metabolic (23.2 +/- 1.3 vs. 30.3 +/- 1.0% in control) but was not altered by respiratory (32.3 +/- 0.6) acidosis. Alkaline phosphatase activity was inhibited by metabolic (402 +/- 16 vs. 471 +/- 15 nmol P.min-1.mg protein-1 in control) but not altered by respiratory (437 +/- 25) acidosis. beta-Glucuronidase activity was stimulated by metabolic (1.02 +/- 0.06 vs. 0.78 +/- 0.05 micrograms phenolphthalein released.bone-1.h-1 in control) but not altered by respiratory (0.73 +/- 0.06) acidosis. Net calcium efflux in control was increased by metabolic (783 +/- 57 vs. 20 +/- 57 nmol.bone-1.48 h-1 in control) and by respiratory (213 +/- 45) acidosis; however, calcium efflux with metabolic was greater than with respiratory acidosis.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals POS0404 VOLUNTARY WHEEL RUNNING MODEL IN MICE TO MECHANICALLY STIMULATE THE ENTHESIS OF THE ACHILLES TENDON

2021 ◽  
Vol 80 (Suppl 1) ◽  
pp. 431.3-431
Author(s):  
A. Briolay ◽  
S. Delplace ◽  
F. Duboeuf ◽  
O. Peyruchaud ◽  
D. Magne ◽  
...  
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Achilles Tendon ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Mechanical Stimulation ◽  
Long Bones ◽  
Serum Samples ◽  
Tissue Specific ◽  
Tissue Extracts

Background:Excessive bone formation in the entheses is one of the features of peripheral spondyloarthritis. Biomechanical stress is proposed to occupy a central place in spondyloarthritis pathophysiology, but the precise molecular and cellular mechanisms underlying the pathological response of the enthesis are still largely unknown [1]. Besides, physical therapy and exercise are recommended as non-pharmacologic therapies for patients. We focused on the effect of exercising on enthesis ossification.Objectives:We aimed to develop and characterize an in vivo model in mice to study the impact of mechanical stimulation on the enthesis of the Achilles tendon.Methods:DBA/1 mice were subjected to voluntary running exercise by the use of activity wheels for two weeks, and compared to mice housed in standard conditions (n=17 per group). The running performances were recorded. mRNAs were extracted from the long bones (flushed tibia and femur) and the ankles’ entheses for real-time PCR analysis. µCT was performed on the femurs. Alkaline phosphatase activity was detected by histology on the anchorage of the Achilles tendon to the calcaneum, and by enzymatic assay in serum samples. Luminex analysis was also conducted on serum samples for Il-6 and Il-8/Kc detection.Results:Free access to the activity wheel resulted in a running exercise of 5.5±0.8 km/day (approximately 80 km in total) at 14.5±0.5 m/min. No effect was detected on the femur architecture by µCT. Sclerostin (Sost) gene expression was monitored as a mechanosensitive marker. Its expression was expectedly reduced by half in entheseal tissues, but no modulation was observed in long bones (Figure 1). Similarly, exercise-induced regulation of Osterix and Runx2 expressions was observed only in enthesis samples. This tissue-specific pattern was also verified for key genes of the sphingosine-1 phosphate metabolic pathway, which we recently implicated in spondyloarthritis pathophysiology [2]. The in situ staining of alkaline phosphatase activity suggested the presence of more positive cells in the anchorage of Achilles tendon of running mice, compared to control ones. However, alkaline phosphatase activity in serum samples and its gene expression in rough tissue extracts were unchanged. No inflammatory response was detected as Il-8/Kc serum levels were similar in the control and the exercising group (59±14 vs 57±14 pg/mL). In addition, Il-6 was not detected in the serum and its expression was very faint and constant in the tissue extracts.Conclusion:This work is still in progress for a more complete characterization of the model. We believe that this experimental design will be useful to study the role of mechanical stimulation specifically in the enthesis and that it can help to better understand the spondyloarthritis pathophysiology.References:[1]Cambré, et al. Nat Commun, 2018; [2] El Jamal, et al. J Bone Miner Res, 2019Figure 1.Expression level of the mechanosensitive gene Sclerostin (Sost). It dropped in response to exercise in entheseal tissues, but not in long bones, revealing a tissue-specific response to mechanical stimulation.Acknowledgements:Société Arthritis R&D (2020)Disclosure of Interests:None declared

scholarly journals Effect of Magnolol on the Function of Osteoblastic MC3T3-E1 Cells

2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Eun Jung Kwak ◽  
Young Soon Lee ◽  
Eun Mi Choi
Keyword(s):  
Alkaline Phosphatase ◽  
Cell Growth ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Collagen Synthesis ◽  
Osteoclast Differentiation ◽  
Osteoblast Function ◽  
Receptor Activator ◽  
Inducing Factors ◽  
Magnolia Officinalis

Objectives.In the present study, the ability of magnolol, a hydroxylated biphenyl compound isolated fromMagnolia officinalis, to stimulate osteoblast function and inhibit the release of bone-resorbing mediators was investigated in osteoblastic MC3T3-E1 cells.Methods.Osteoblast function was measured by cell growth, alkaline phosphatase activity, collagen synthesis, and mineralization. Glutathione content was also measured in the cells. Bone-resorbing cytokines, receptor activator of nuclear factor-κB ligand (RANKL), TNF-α, and IL-6 were measured with an enzyme immunoassay system.Results.Magnolol caused a significant elevation of cell growth, alkaline phosphatase activity, collagen synthesis, mineralization, and glutathione content in the cells (P<0.05). Skeletal turnover is orchestrated by a complex network of regulatory factors. Among cytokines, RANKL, TNF-α, and IL-6 were found to be key osteoclastogenetic molecules produced by osteoblasts. Magnolol significantly (P<0.05) decreased the production of osteoclast differentiation inducing factors such as RANKL, TNF-α, and IL-6 in the presence of antimycin A, which inhibits mitochondrial electron transport and has been used as an ROS generator.Conclusion.Magnolol might be a candidate as an agent for the prevention of bone disorders such as osteoporosis.

Influence of osteoclasts and osteoclast-like cells on osteoblast alkaline phosphatase activity and collagen synthesis

2009 ◽  
Vol 9 (8) ◽  
pp. 1167-1178 ◽  
Author(s):  
Rachelle J. Sells Galvin ◽  
James W. Cullison ◽  
Louis V. Avioli ◽  
Philip A. Osdoby
Keyword(s):  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Collagen Synthesis
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