Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2022 Nov 4;23(21):13554.
doi: 10.3390/ijms232113554.

Hemizygous Granzyme A Mice Expressing the hSOD1G93A Transgene Show Slightly Extended Lifespan

Laura Moreno-Martinez  1   2   3   4 , Llipsy Santiago  3   5 , Miriam de la Torre  1   2   3   4 , Ana Cristina Calvo  1   2   3   4 , Julián Pardo  3   5   6 , Rosario Osta  1   2   3   4
Affiliations

Hemizygous Granzyme A Mice Expressing the hSOD1G93A Transgene Show Slightly Extended Lifespan

Laura Moreno-Martinez et al. Int J Mol Sci. .

Abstract

Granzyme A (gzmA), a serine protease involved in the modulation of the inflammatory immune response, is found at an elevated level in the serum from ALS patients. However, the influence of gzmA on the progression of ALS remains unclear. The aim of our work was to assess whether the absence of gzmA in an ALS murine model could help slow down the progression of the disease. Homozygous and hemizygous gzmA-deficient mice expressing the hSOD1G93A transgene were generated, and survival of these mice was monitored. Subsequently, gene and protein expression of inflammatory and oxidative stress markers was measured in the spinal cord and quadriceps of these mice. We observed the longest lifespan in gzmA+/- mice. GzmA gene and protein expression was downregulated in the spinal cord and serum from gmzA+/- mice, confirming that the increased survival of hemizygous mice is correlated with lower levels of gzmA. In addition, mRNA and protein levels of glutathione reductase (GSR), involved in oxidative stress, were found downregulated in the spinal cord and quadriceps of gmzA+/- mice, together with lower IL-1β and IL-6 mRNA levels in hemyzigous mice. In summary, our findings indicate for the first time that reduced levels, but not the absence, of gzmA could slightly ameliorate the disease progression in this animal model.

Keywords: SOD1G93A mouse; amyotrophic lateral sclerosis; glutathione reductase; granzyme A; inflammation.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Figures

Figure 1
Figure 1
Survival analysis of SOD1G93A mice varying in gzmA genotype. Survival analysis was performed with mice showing the three gzmA genotypes (n = 34 gzmA−/−, n = 49 gzmA+/− and n = 19 gzmA+/+). Survival curve showing statistically significant differences in the lifespan between gzmA+/− and gzmA+/+ mice (p = 0.022).
Figure 2
Figure 2
hSOD1G93A expression in spinal cord and quadriceps of gzmA+/+, gzmA+/− and gzmA−/− mice involved in the survival assay. (a) Expression of hSOD1G93A was quantified in both tissues from 10 gmzA+/+, 10 gzmA+/− and 10 gzmA−/− mice. No differences between genotypes were found (p > 0.05). (b) Correlations between hSOD1G93A expression and survival time were not statistically significant (r2 = 0.0000884, p = 0.979 in spinal cord, r2 = 0.0649, p = 0.508 in quadriceps).
Figure 3
Figure 3
GzmA gene and protein expression in serum, spinal cord and quadriceps from SOD1G93A mice. (a) Gzma gene expression was analyzed in spinal cord and quadriceps of 16 SOD1+/− mice from the survival assay, including gzmA+/+(n = 6), gzmA+/− (n = 6) and gzmA−/− (n = 4) mice. Gzma levels were found downregulated in spinal cord from gzmA+/− mice compared to the levels found in gzmA+/+ mice (p = 0.001). Similar tendency was observed in quadricep tissue (p = 0.076). (b) Serum gzmA was measured in 33 SOD1+/− mice at 90 days old, including gzmA+/+ (n = 7), gzmA+/− (n = 14) and gzmA−/− (n = 12) genotypes. Serum gzmA levels were found reduced in gzmA+/− mice (p = 0.005). Bars represent mean ± standard error of the mean. ** p < 0.01, ^ p < 0.10.
Figure 4
Figure 4
Il1b, Il6, Tnfa and Gsr gene expression in spinal cord (a) and quadriceps (b) from SOD1G93A mice. Il1b, Il6, Tnfa and Gsr gene expression was analyzed in spinal cord and quadriceps of 30 SOD1+/− mice, including gzmA+/+ (n = 10), gzmA+/− (n = 10) and gzmA−/− (n = 10) mice from the survival assay. (a) Il1b, Il6 and Gsr levels were found reduced in spinal cord from gzmA+/− mice. (b) Il1b and Tnfa levels were reduced in quadriceps from gzmA+/− mice. Bars represent mean ± standard error of the mean. ** p < 0.01, * p < 0.05.
Figure 5
Figure 5
IL1B, IL6 and GSR protein expression in spinal cord (a) and quadriceps (b) from SOD1G93A mice from the survival assay. IL1Β, IL6 and GSR protein expression was analyzed in spinal cord and quadriceps of 12 SOD1+/− mice, including gzmA+/+(n = 4), gzmA+/− (n = 4) and gzmA−/− (n = 4) mice. GSR was found at lower levels in hemizygous mice compared to both homozygous mice. Levels of IL6 in spinal cord and IL1B in quadriceps were reduced in gzmA+/+ and gzmA+/− mice. ** p < 0.01, * p < 0.05.
Figure 6
Figure 6
Scheme of crossings to generate homozygous and hemizygous gzmA mice. Hemizygous SOD1G93A males (SOD1+/−, gzmA+/+) were crossed with C57BL/6 females deficient in gzmA (SOD1−/−, gzmA−/−) to obtain hemizygous gzmA mice. Afterwards, we crossed hemizygous SOD1G93A and gzmA mice (SOD1+/−, gzmA+/−) to generate littermate mice with the three gzmA genotypes with the same genetic background.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Hardiman O., Al-Chalabi A., Chio A., Corr E.M., Logroscino G., Robberecht W., Shaw P.J., Simmons Z., van den Berg L.H. Amyotrophic Lateral Sclerosis. Nat. Rev. Dis. Primers. 2017;3:17071. doi: 10.1038/nrdp.2017.71. - DOI - PubMed
    1. Phani S., Re D.B., Przedborski S. The Role of the Innate Immune System in ALS. Front. Pharmacol. 2012;3:150. doi: 10.3389/fphar.2012.00150. - DOI - PMC - PubMed
    1. Ransohoff R.M. How Neuroinflammation Contributes to Neurodegeneration. Science. 2016;353:777–783. doi: 10.1126/science.aag2590. - DOI - PubMed
    1. Liu J., Wang F. Role of Neuroinflammation in Amyotrophic Lateral Sclerosis: Cellular Mechanisms and Therapeutic Implications. Front. Immunol. 2017;8:1005. doi: 10.3389/fimmu.2017.01005. - DOI - PMC - PubMed
    1. Rodrigues Lima-Junior J., Sulzer D., Lindestam Arlehamn C.S., Sette A. The Role of Immune-Mediated Alterations and Disorders in ALS Disease. Hum. Immunol. 2021;82:155–161. doi: 10.1016/j.humimm.2021.01.017. - DOI - PMC - PubMed