Indomethacin augments lipopolysaccharide-induced expression of inflammatory molecules in the mouse brain

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Biochemistry, Biophysics and Molecular Biology

Main article text

 

Introduction

Materials and Methods

Animals, LPS administration and drug treatment

Wes™ capillary-based protein electrophoresis

Real time RT-PCR

Statistical analyses

Results

Effect of LPS on expression of proinflammatory and anti-inflammatory molecules in the brain

Effect of indomethacin on Il10, Il1b, Tnf and Nos2 mRNA expression

Effects of treatment with indomethacin on LPS-induced changes of Il10, Il1b, Tnf and Nos2 mRNA expression

Effects of treatment with indomethacin on LPS-induced changes of iNOS protein expression

Effects of treatment with minocycline alone or with indomethacin on LPS-induced changes of Il1b, Tnf and Nos2 mRNA expression

Discussion

Conclusions

Supplemental Information

Relative expression of Il10, Il1b, Tnf and Nos2 mRNA in the brain of control (vehicle-only ), LPS-inoculated vehicle-treated and LPS-inoculated indomethacin-treated at 4 h post LPS/vehicle inoculation.

DOI: 10.7717/peerj.10391/supp-1

Expression of Iba-1 protein in the brain of control (vehicle-only) and LPS-inoculated mice at 4 h post LPS/vehicle inoculation.

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Relative expression of Il10, Il1b, Tnf and Nos2 mRNA in the brain of control (vehicle-only ), LPS-inoculated vehicle-treated and LPS-inoculated indomethacin plus minocycline-treated at 4 h post LPS/vehicle inoculation.

Relative expression of Il10, Il1b, Tnf and Nos2 mRNA in the brain of control (vehicle-only ), LPS-inoculated vehicle-treated and LPS-inoculated indomethacin plus minocycline-treated at 4 h post LPS/vehicle inoculation.

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Relative expression of Il1b, Tnf and Nos2 mRNA in the brain of control (vehicle-only) and LPS-inoculated mice at 4 h post LPS/vehicle inoculation.

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Expression of iNOS protein in the brain of control (vehicle-only), LPS-inoculated vehicle-treated and LPS-inoculated indomethacin-treated at 4 h post LPS/vehicle inoculation.

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Calculations for Figure 1.

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Relative expression of Il10, Il1b, Tnf and Nos2 mRNA in the brain of control (vehicle-only ), LPS-inoculated vehicle-treated and LPS-inoculated minocycline-treated at 4 h post LPS/vehicle inoculation.

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Relative expression of Il1b, Tnf and Nos2 mRNA in the brain of control (vehicle-only) and indomethacin-treated mice at 4 h post treatement.

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Virtual-blot like image similar to traditional results obtained from electropherograms for Figure 1.

Lane 1 and 2 is for actin, lane 3 and 4 is for Iba-1

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Electropherograms of actin for Figure 1.

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Electropherograms of Iba-1for Figure 1.

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Illustration of Wes data analysis.

Data analysis using ratio the between of the areas of the resulting electropherograms for Iba-1 and β-actin generated during the Wes capillary-based protein electrophoresis.

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Effect of inoculation of BALB/c mice with lipopolysaccharide (LPS) on Iba-1 protein expression.

Protein expression of Iba-1 in the brain of control (vehicle-only) and LPS-inoculated mice at 4 h post LPS/vehicle inoculation. Each box and whiskers represents the median and interquartile range of the values obtained from five animals. Statistically significant difference with vehicle-treated control animals: **p < 0.01 (Mann–Whitney U test).

DOI: 10.7717/peerj.10391/supp-13

Effect of inoculation of BALB/c mice with lipopolysaccharide (LPS) on Il10, Il1b, Tnf and Nos2 mRNA expression.

Relative expression of (A) Il10, (B) Il1b, (C) Tnf and (D) Nos2 mRNA in the brain of control (vehicle-only) and LPS-inoculated mice at 4 h post LPS/vehicle inoculation. Each bar or box and whiskers represents the mean ± S.E.M (A and B) or median and interquartile range (C), respectively, of the values obtained from eight animals. Statistically significant difference with vehicle-treated control animals: ***p < 0.01 (Student’s t test for Il10, Il1b and Tnf or Mann–Whitney U test for Nos2).

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Supplemental results.

DOI: 10.7717/peerj.10391/supp-15

Additional Information and Declarations

Competing Interests

The authors declare that they have no competing interests.

Author Contributions

Mona Yasin Mohamed performed the experiments, analyzed the data, prepared figures and/or tables, and approved the final draft.

Willias Masocha conceived and designed the experiments, analyzed the data, prepared figures and/or tables, authored or reviewed drafts of the paper, and approved the final draft.

Animal Ethics

The following information was supplied relating to ethical approvals (i.e., approving body and any reference numbers):

The Ethical Committee for Use of Laboratory Animals in Teaching and Research of the Health Sciences of Kuwait University approved this research.

Data Availability

The following information was supplied regarding data availability:

The protein expression raw data and real-time PCR raw data are available in the Supplemental Files.

The protein expression was done using the ProteinSimple’s WesTM Capillary-Based Protein Electrophoresis; thus, the raw data comes as electropherograms from which virtual-blot-like images similar to traditional results as well as a Supplemental File showing the height and area under the electropherograms are generated.

Funding

This study was supported by grant YP 01/18 from Kuwait University Research Sector and the Core Research Facility KU project # OMICSRU/RCF project SRUL02/13 for the real-time PCR equipment. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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