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. 2024 Oct 10;46(10):11282-11295.
doi: 10.3390/cimb46100670.

TLR4 Downregulation Identifies High-Risk HPV Infection and Integration in H-SIL and Squamous Cell Carcinomas of the Uterine Cervix

Angela Santoro  1   2 , Giuseppe Angelico  3 , Damiano Arciuolo  1 , Giulia Scaglione  1 , Belen Padial Urtueta  1 , Gabriella Aquino  4 , Noemy Starita  5 , Maria Lina Tornesello  5 , Rosalia Anna Rega  6 , Maria Carmela Pedicillo  7 , Manuel Mazzucchelli  8 , Ilenia Sara De Stefano  7 , Rosanna Zamparese  9 , Giuseppina Campisi  10 , Giorgio Mori  7 , Gian Franco Zannoni  1   2 , Giuseppe Pannone  7
Affiliations

TLR4 Downregulation Identifies High-Risk HPV Infection and Integration in H-SIL and Squamous Cell Carcinomas of the Uterine Cervix

Angela Santoro et al. Curr Issues Mol Biol. .

Abstract

Growing scientific evidence suggests a link between the expression of toll-like receptor 4 (TLR4) and cervical cancer carcinogenesis. Specifically, a close relation between TLR4 expression and FIGO stage, lymph node metastases, and tumor size has been reported in cervical cancer. In the present study, we aimed to evaluate the relationship between TLR4 expression levels and human papillomavirus (HPV) infection and/or high-risk (hr) HPV integration status in patients with a histological diagnosis of high-grade squamous intraepithelial lesion (H-SIL), and squamous cell carcinoma (SCC) of the uterine cervix. Sixty biopsies of cervical neoplasia, comprising H-SIL (n = 20) and SCC (n = 40), were evaluated for TLR4 expression by immunohistochemistry. All samples were positive for high-risk HPV as confirmed by in situ hybridization (ISH) and broad-spectrum PCR followed by Sanger sequencing analysis. The intensity of TLR4 staining was higher in tissues negative for intraepithelial lesion or malignancy (NILM) than in H-SIL, and further reduced in SCC. Moreover, statistically significant differences have been observed in the percentage of TLR4 expression between NILM and H-SIL and between H-SIL and SCC, with higher percentages of expression in H-SIL than in SCC. Our results showed a significant downregulation of TLR4 in HPV-related H-SIL and SCC, compared to NILM. These data support the hypothesis that TLR4 expression is suppressed in HPV-driven oncogenesis.

Keywords: HPV; TLR4; cervical cancer; immunotherapy; p16.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
TLR4 expression intensity and percentage. The analysis of TLR4 SI in NILM, H-SIL, and SCC samples showed a statistically significant difference between the 3 groups.
Figure 2
Figure 2
(A) NILM showing negative immunostaining for TLR4. (B) H-SIL showing diffuse and intense immunostaining for TLR4.
Figure 3
Figure 3
TLR4 staining patterns. (A,B) Double immunostaining for p16 (DAB–brown) and TLR-4 (Fast Red) in SCC. With double immunostaining, p16 staining was distributed at the periphery of the tumoral nests while TLR4 was more evident centrally, in more differentiated tumoral cells. (C) A case of H-SIL showing low TLR-4 expression (downregulation) is depicted. (D) A case of SCC showing moderate TLR4 expression.
Figure 4
Figure 4
Immunohistochemical expression of CK19. The analysis of percentages of immunohistochemical expression of CK19 showed a statistically significative difference between normal epithelium, H-SIL, and SCC (p < 0.05; ANOVA).
Figure 5
Figure 5
Correlation expression of TLR4 and CK19. Inverse statistically significant correlation between TLR4-SI and CK19 percentage of expressions (R = −0.4906; p = 0.0001, ANOVA), with CK19 upregulation and TLR4 downregulation in the cervical carcinogenetic process.
Figure 6
Figure 6
CK19 upregulation and TLR4 downregulation in the cervical carcinogenetic process. (A) Immunohistochemical expression of CK19 SCC; (B) immunohistochemical expression of TLR4 in the same case of SCC.
Figure 7
Figure 7
Correlation between intensity of TLR-4 expression and the degree of inflammatory infiltration. A statistically significant correlation between TLR4 downregulation and inflammatory tumoral microenvironment has been demonstrated (R = 0.5341; p = 0.001, ANOVA).
See this image and copyright information in PMC

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