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. 2023 Jan 17:81:ftad004.
doi: 10.1093/femspd/ftad004.

Murine herpesvirus-68-related growth factors treatment correlates with decrease of p53 and HIF-1α protein levels

Miroslava Supolikova  1 , Eva Novakova  1   2 , Karin Donatova  2 , Petra Olejnikova  3 , Martina Labudova  1   2
Affiliations

Murine herpesvirus-68-related growth factors treatment correlates with decrease of p53 and HIF-1α protein levels

Miroslava Supolikova et al. Pathog Dis. .

Abstract

Murine herpesvirus 68 (MHV-68) belongs to the subfamily Gammaherpesvirinae of the family Herpesviridae. This exceptional murine herpesvirus is an excellent model for the study of human gammaherpesvirus infections. Cells infected with MHV-68 under nonpermissive conditions for viral replication produce substances designated as MHV-68 growth factors (MHGF-68), that can cause transformation of the cells, or on the other side, turn transformed cells into normal. It was already proposed, that the MHGF-68 fractions cause transformation, disruption of the cytoskeleton and slower growth of the tumors in nude mice. Here, we examined newly extracted fractions of MHGF-68 designated as F5 and F8. Both fractions proved to inhibit the growth of the spheroids and also tumours induced in nude mice. What more, the fractions caused the decrease of the protein levels of wt p53 and HIF-1α. Decreased levels of p53 and HIF-1α activity leads to decreased vascularization, slower tumour growth, and lower adaptation to hypoxic conditions. This would propose MHGF-68 fractions, or their human herpesvirus equivalents, as a potential anticancer drugs in combined chemotherapy.

Keywords: HIF-1α; MHGF-68; MHV-68; growth factors; mdm2; p53.

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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.
The presence of MHGF-68 correlates with low levels of p53. (A) Western blot of Hepa1c1c7 cell monolayers under normoxic (NO) and hypoxic conditions. The cells were cultivated in hypoxic atmosphere (1% hypoxia) or treated with CoCl2 to induce hypoxia. Cells were treated with MHGF-68 fractions F5 and F8 and control fraction F7. Protein p53 and actin levels are shown. The MHGF-68 fractions inhibit the spheroid growth. (B) Hepa1c1c7 cell line spheroid photographs on days 3, 9, and 11. Magnification 50x. (C) Area of Hepa1c1c7 spheroids as determined by Image J software in pixels. (D) Distribution of spheroid sizes at the 11th day of the treatment with cross-representing average value. The experiments were done in three independent experiments. Statistical significance was evaluated by Student’s t-test. Where * is P < .05; ** is P < .01; and *** is P < .001.
Figure 2.
Figure 2.
The MHGF-68 fractions inhibit the tumour growth. (A) Tumour growth presented as tumour volume in mm3 observed for 16 days after treatment. (B) Boxed graph showing the distribution of tumour volumes. The graph shows the volumes obtained at day 16 measured on live mice. (C) Boxed graph showing the distribution of tumour weights on the 16th day, with cross-representing average value and circles inside and extreme values. Statistical significance was evaluated by Student’s t-test. Where * is P < .05; ** is P < .01; and *** is P < .001. (D) Representative xenografts of control, F5- and F8-treated mice. The treated groups were compared to the control groups (A–C).
Figure 3.
Figure 3.
Western blot analysis of p53 and MDM2 expression. (A) Western blot analysis of the tumours from untreated or MHGF-68-treated mice. The level of total p53, serine 15 phosphorylated/active p53, and MDM2 was evaluated by Image J software. Graph of (B) p53, (C) serine 15 phosphorylated p53, and (D) MDM2 band intensity measured by Image J software. The band intensities were normalized to the actin. (E) The delta Ct values of P53 expression levels in treated and un-treated tumours. The treated tumours have higher Ct values than the controls as they have lower levels of mRNA. The percentage is compared to the control. The experiments were done in three independent experiments. Statistical significance was evaluated by Student’s t-test. Where * is P < .05; ** is P < .01; and *** is P < .001.
Figure 4.
Figure 4.
Immunohistochemistry of tumours treated with MHGF-68 fraction.
Figure 5.
Figure 5.
Western blot analysis of HIF-1α and MMP13 expression. (A) Western blot analysis of the tumours from untreated or MHGF-68-treated mice. Graph of (B) HIF-1α and (C) MMP13 band intensity measured by Image J software. The band intensities were normalized to the actin. The percentage is compared to the control. The experiments were done in independent experiments. Statistical significance was evaluated by Student’s t-test. Where * is P < .05; ** is P < .01; and *** is P < .001.
Figure 6.
Figure 6.
Diagram of proposed hypothetical MHGF-68 mechanism of action. Viral IRF1/MHGF-68 fraction interacts with ATM kinase, blocking its activity followed by reduction of p53 phosphorylation at the serine 15 residue. The inhibition of phosphorylation leads to the higher p53 ubiquitination and decreased levels of p53. Viral IRF4/MHGF-68 fraction interacts with MDM2 and suppresses its ubiquitination. Higher levels of MDM2 then lead to enhanced p53 ubiquitination and decreased levels of p53.
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References

    1. Abràmoff MD, Magalhães PJ, Ram SJ. Image processing with imageJ. Biophotonics Int. 2004;11:36–42.
    1. Ala-Aho R, Grénman R, Seth Pet al. . Adenoviral delivery of p53 gene suppresses expression of collagenase-3 (MMP-13) in squamous carcinoma cells. Oncogene. 2002;21:1187–95. - PubMed
    1. An WG, Kanekal M, Simon MCet al. . Stabilization of wild-type p53 by hypoxia-inducible factor 1α. Nature. 1998;392:405–8. - PubMed
    1. Bergers G, Javaherian K, Lo KMet al. . Effects of angiogenesis inhibitors on multistage carcinogenesis in mice. Science. 1999;284:808–12. - PubMed
    1. Boehm T, Folkman J, Browder Tet al. . Antiangiogenic therapy of experimental cancer does not induce acquired drug resistance. Nature. 1997;390:404–7. - PubMed

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