Optimal parameters for the enhancement of human osteoblast-like cell proliferation in vitro via shear stress induced by high-frequency mechanical vibration

Introduction: Biomechanical stimulation of cultured human osteoblast-like cells, which is based on controlled mechanical vibration, has been previously indicated, but the exact mechanical parameters that are effective for cells' proliferation enhancement are still elusive due to the lack of direct data recordings from the stimulated cells in culture. Therefore, we developed a low friction tunable system that enables recording of a narrow range of mechanical parameters, above the infrasonic spectrum, that applied uniformly to human osteoblast-like cells in monolayer culture, aiming to identify a range of mechanical parameters that are effective to enhance osteoblast proliferation in vitro. Methods: Human osteoblast-like cells in explant monolayer culture samples were exposed to mechanical vibration in the 10-70Hz range of frequencies for two minutes, in four 24 hours intervals. Cell numbers in culture, cellular alkaline phosphatase activity (a marker of cell maturation), and lactate dehydrogenase activity in culture media (representing cell death) were measured after the mechanical stimulation protocol application and compared statistically to the control cell cultures kept in static conditions. The cell proliferation was deduced from cell number in culture and cell death measurements. Results: We found that 50-70 Hz of vibration frequency protocol (10-30 μm of maximal displacement amplitude, 0.03g of peak-to-peak acceleration) is optimal for enhancing cells' proliferation(p<0.05), with a parallel decrease of their maturation (p<0.01). Discussion: We detected the optimal mechanical parameters of excitation protocol for induction of osteoblast proliferation in vitro by a mechanical platform, which can be used as a standardized method in the research of mechanotransduction in human osteoblast.

Download Full-text

Determination of biophysical parameters for enhancement of human osteoblast proliferation by mechanical vibration in the infrasonic frequency range

10.1101/448225 ◽

2018 ◽

Author(s):

Rosenberg Nahum ◽

Halevi Politch Jacob ◽

Rosenberg Orit ◽

Abramovich Haim

Keyword(s):

Mechanical Stimulation ◽

Culture Media ◽

Mechanical Vibration ◽

Mechanical Parameters ◽

Human Osteoblast ◽

Osteoblast Proliferation ◽

Biophysical Parameters ◽

Proliferation In Vitro ◽

Stimulation Of

AbstractExperimental methods for studying an enhancement of osteoblast proliferation in vitro provide tools for the research of biochemical processes involved in bone turnover in vivo. Some of the current methods used for this purpose are based on the ability of the osteoblasts to enhance proliferation by mechanical stimulation. We describe an experimental approach of biomechanical stimulation of cultured human osteoblast-like cells by vibration. This method is based on the specially designed controlled vibration setup that consists of an electric actuator, with horizontally mounted well plate containing cell cultures. Previously this method found to be effective to enhance cell proliferation, but the exact mechanical parameters of effective vibration were elusive. The current low friction system for mechanical stimulation of osteoblast-like cells in vitro provides recording of narrow range mechanical parameters in the infrasonic spectrum.We exposed human osteoblast-like cells in explant monolayer culture to mechanical vibration in the 10-70Hz range of frequencies and found that 50-70 Hz of vibration frequency is optimal for inducing osteoblast proliferation that was deduced from interrelation between unchanged cell number in culture samples with significant decrease in cell death rate (decreased LDH activity in culture media, p<.05) and with parallel decrease of their maturation level (p<.01).In this report we determined the optimal mechanical parameters and excitation protocol for induction of osteoblast proliferation in vitro by using a tunable and versatile mechanical platform, which can be used in the research of cell mechanotransduction.

Download Full-text

Age-Related Changes in the Effects of Serum from Lean and Obese Pigs on Myogenic Cell Proliferation in Vitro

Journal of Animal Science ◽

10.2527/jas1982.544763x ◽

1982 ◽

Vol 54 (4) ◽

pp. 763-768 ◽

Cited By ~ 3

Author(s):

Ronald E. Allen ◽

Gail Robinson ◽

Matthew J. Parsons ◽

Robert A. Merkel ◽

William T. Magee

Keyword(s):

Cell Proliferation ◽

Myogenic Cell ◽

Age Related ◽

Proliferation In Vitro ◽

Age Related Changes

Download Full-text

The P-MAPA immunomodulator partially prevents apoptosis induced by Zika virus infection in THP-1 cells

Current Pharmaceutical Biotechnology ◽

10.2174/1389201021666200602140005 ◽

2020 ◽

Vol 21 ◽

Cited By ~ 1

Author(s):

Morganna C. Lima ◽

Elisa A. N. Azevedo ◽

Clarice N. L. de Morais ◽

Larissa I. O. de Sousa ◽

Bruno M. Carvalho ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Death ◽

Virus Infection ◽

Virus Replication ◽

Zika Virus ◽

Mammalian Cells ◽

Caspase 3 ◽

Neurological Complications ◽

Zika Virus Infection

Background: Zika virus is an emerging arbovirus of global importance. ZIKV infection is associated with a range of neurological complications such as the Congenital Zika Syndrome and Guillain Barré Syndrome. Despite the magnitude of recent outbreaks, there is no specific therapy to prevent or to alleviate disease pathology. Objective: To investigate the role of P-MAPA immunomodulator in Zika-infected THP-1 cells. Methods: THP-1 cells were subjected at Zika virus infection (Multiplicity of Infection = 0.5) followed by treatment with P-MAPA for until 96 hours post-infection. After that, the cell death was analyzed by annexin+/ PI+ and caspase 3/ 7+ staining by flow cytometry. In addition, the virus replication and cell proliferation were accessed by RT-qPCR and Ki67 staining, respectively. Results: We demonstrate that P-MAPA in vitro treatment significantly reduces Zika virus-induced cell death and caspase-3/7 activation on THP-1 infected cells, albeit it has no role in virus replication and cell proliferation. Conclusions: Our study reveals that P-MAPA seems to be a satisfactory alternative to inhibits the effects of Zika virus infection in mammalian cells.

Download Full-text

Single-chain Antibody Against Reg4 Suppresses Gastric Cancer Cell Growth and Enhances 5-FU-induced Cell Death in vitro

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520619666181122104720 ◽

2019 ◽

Vol 19 (5) ◽

pp. 610-619 ◽

Cited By ~ 3

Author(s):

Xue-Qing Zhang ◽

Lu-Ting Yu ◽

Pei Du ◽

Tian-Qi Yin ◽

Zhi-Yuan Zhang ◽

...

Keyword(s):

Gastric Cancer ◽

Cell Proliferation ◽

Cell Death ◽

Cancer Cell ◽

Gastric Cancer Cell ◽

Single Chain ◽

Single Chain Antibody ◽

Ags Cells ◽

Thiazolyl Blue Tetrazolium Bromide

Background:Regenerating islet-derived gene family member 4 (Reg4), a well-investigated growth factor in the regenerative pancreas, has recently been reported to be highly associated with a majority of gastrointestinal cancers. Pathological hyper-expression or artificial over-expression of Reg4 causes acceleration of tumor growth, migration, and resistance to chemotherapeutic 5-Fluorouracil (5-FU). Until now, no method has been successfully established for eliminating the effects of Reg4 protein.Methods:This study reports the production of an engineered immunoglobin, a single-chain variable fragment (scFv-Reg4), to specifically bind Reg4 and block the bioactivity. The complementary-determining regions (CDRs) against Reg4 were assigned using MOE and ZDOCK servers. The binding affinity (KD) was determined by bio-layer interferometry (BLI). MKN45 and AGS cell proliferation was determined by Thiazolyl blue tetrazolium bromide (MTT) method and the cell apoptosis was detected by flow cytometry assay.Results:The KD of scFv-Reg4 to Reg4 was determined to be 1.91×10-8. In MKN45 and AGS cell lines, scFv- Reg4 depressed Reg4-stimulated cell proliferation and the inhibitory rates were 27.7±1.5% and 17.3±2.6%, respectively. Furthermore, scFv significantly enhanced 5-FU-induced cell death, from 23.0±1.0% to 28.4±1.2% in MKN45 and 28.2±0.7% to 36.6±0.6% in AGS cells. Treatment with scFv alone could lyse cancer cells to a certain extent, but no significance has been observed.Conclusion:The single-chain antibody (scFv-Reg4) significantly inhibited gastric cancer cell proliferation and synergistically enhanced the lethal effect of 5-FU. Thus, traditional chemo-/radio- therapeutics supplemented with scFv-Reg4 may provide advances in the strategy for gastrointestinal cancer treatment.

Download Full-text