12 research outputs found
Collaborative study of thresholds for mutagens: proposal of a typical protocol for detection of hormetic responses in cytotoxicity tests
Abstract Background According to the linear no-threshold model (LNT), even the smallest amount of radiation is hazardous. Although the LNT is not based on solid data, this hypothesis has been applied to mutagens and carcinogens. As a result, it has been postulated that there are no thresholds for these chemicals. To demonstrate negativity by experiments is practically impossible, because negative data may leave behind the possibility that additional data might make the resolution power high enough to change negativity to positivity. Furthermore, additional data collection may be endless and we may be trapped in agnosticism. When hormesis is established, in which biological responses are higher at low-doses and lower at high-doses than the control, thresholds could be established between the low- and high-doses. Before examination of thresholds in chemical mutagenesis, hormetic responses in cytotoxicity were tested using cultured mammalian cells. Method Human cells (HeLa S3 and TK6) or Chinese hamster cells (CHL/IU) were cultured in 96-well plates and treated with mitomycin C (MMC) or ethyl methanesulfonate (EMS) at various dose levels and optical density was measured after addition of a reagent to detect cellular activity. In hormetic responses, data might fluctuate to and fro; therefore, experimental conditions were examined from various aspects to eliminate confounding factors including cell numbers, detection time, the edge effect of 96-well plates, and measurement time after addition of the reagent for detection. Results The dose response relationship was never linear. Cellular activities after treatment with MMC or EMS were generally higher at lower doses levels and lower at higher doses than the control, showing hormesis and allowing the establishment of thresholds. Dose response curves sometimes showed two or three peaks, probably reflecting different cellular responses. Conclusion Hormetic responses in cytotoxicity tests were observed and thresholds could be established. Based on the results of this investigation, we put forward a tentative protocol to detect chemical hormesis in cytotoxicity tests, i.e., inoculate 2000 cells per well, add various doses of a test chemical 48 h after inoculation, add a detection dye 10 h after treatment, and measure optical density 2 h after addition of the reagent for detection
Construction and Characterization of Protein-Encapsulated Electrospun Fibermats Prepared from a Silica/Poly(γ-glutamate) Hybrid
Protein-encapsulated fibermats are
an attractive platform for protein-based
bioactive materials. However, the choice of methods is still limited
and not applicable to a wide range of proteins. In this study, we
studied new polymeric materials for constructing protein-encapsulated
fibermats, in which protein molecules are encapsulated within the
nanofibers of fibermats without causing deleterious changes to protein
structure or function. We constructed a protein-encapsulated fibermat
using the poly(γ-glutamate) (PGA)/(3-glycidyloxypropyl)-trimethoxysilane
(GPTMS) hybrid as a precursor for electrospinning. Because the PGA/GPTMS
hybrid is water-soluble, protein molecules can be added to the precursor
in an aqueous solution, significantly enhancing protein stability.
Polycondensation during electrospinning (in-flight polycondensation)
makes the obtained fibermats water-insoluble, which stabilizes the
fibermat structure such that it is resistant to degradation in aqueous
buffer. The molecular structure of the PGA/GPTMS hybrid gives rise
to unique molecular permeability, which alters the selectivity and
specificity of biochemical reactions involving the encapsulated enzymes;
lower molecular-weight (MW) substrates can permeate the nanofibers,
promoting enzyme activity, but higher MW substrates such as inhibitor
peptides cannot permeate the nanofibers, suppressing enzyme activity.
We present an effective method of encapsulating bioactive molecules
while maintaining their structure and function, increasing the versatility
of electrospun fibermats for constructing various bioactive materials