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We present the findings of assessing the expression levels of extracellular TLR2 and TLR4 and intracellular TLR3 and TLR8 correlating with the severity of clinical manifestations of HPV infection. A total of 199 women took part in a single-center prospective comparative research study on TLR2, TLR3, TLR4 and TLR8 expression in HPV-related cervical lesions. TLRs' mRNA expression was analyzed using real-time reverse transcription polymerase chain reaction (RT-PCR). Our results indicate the potential significance of TLR3, TLR4 and TLR8 in responding to HPV infection and its progression to SILs and CC, highlighting the importance of HPV polyinfection in relation to TLR4 and TLR8. [ABSTRACT FROM AUTHOR]

Objective In this study, we established a Sprague-Dawley rat model of vulvar squamous intraepithelial lesions and investigated the impact of focused ultrasound on the expression of hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF) and mutant type p53 (mtp53) in the vulvar skin of rats with low-grade squamous intraepithelial lesions (LSIL).Materials and methods The vulvar skin of 60 rats was treated with dimethylbenzanthracene (DMBA) and mechanical irritation three times a week for 14 weeks. Rats with LSIL were randomly allocated into the experimental group or the control group. The experimental group was treated with focused ultrasound, while the control group received sham treatment.Results After 14 weeks treatment of DMBA combined with mechanical irritation, LSIL were observed in 44 (73.33%) rats, and high-grade squamous intraepithelial lesions (HSIL) were observed in 14 (23.33%) rats. 90.91% (20/22) of rats showed normal pathology and 9.09% (2/22) of rats exhibited LSIL in the experimental group at four weeks after focused ultrasound treatment. 22.73% (5/22) of rats exhibited LSIL, 77.27% (17/22) of rats progressed to HSIL in the control group. Compared with the control-group rats, the levels of HIF-1α, VEGF and mtp53 were significantly decreased in experimental-group rats (p

The lack of human‐derived in vitro models that recapitulate cervical pre‐cancerous lesions has been the bottleneck in researching human papillomavirus (HPV) infection‐associated pre‐cancerous lesions and cancers for a long time. Here, a long‐term 3D organoid culture protocol for high‐grade squamous intraepithelial lesions and cervical squamous cell carcinoma that stably recapitulates the two tissues of origin is described. Originating from human‐derived samples, a small biobank of cervical pre‐tumoroids and tumoroids that faithfully retains genomic and transcriptomic characteristics as well as the causative HPV genome is established. Cervical pre‐tumoroids and tumoroids show differential responses to common chemotherapeutic agents and grow differently as xenografts in mice. By coculture organoid models with peripheral blood immune cells (PBMCs) stimulated by HPV antigenic peptides, it is illustrated that both organoid models respond differently to immunized PBMCs, supporting organoids as reliable and powerful tools for studying virus‐specific T‐cell responses and screening therapeutic HPV vaccines. In this study, a model of cervical pre‐cancerous lesions containing HPV is established for the first time, overcoming the bottleneck of the current model of human cervical pre‐cancerous lesions. This study establishes an experimental platform and biobanks for in vitro mechanistic research, therapeutic vaccine screening, and personalized treatment for HPV‐related cervical diseases. [ABSTRACT FROM AUTHOR]

Introduction: Human papilloma virus (HPV) is the most common sexually transmitted infection worldwide. Cervicovaginal microbiota plays an important role in HPV infection and is associated with the development of squamous intraepithelial lesions (SIL). The natural history of cervical cancer involves reversible changes in the cervical tissue from a normal state, in which no neoplastic changes are detected in the squamous epithelium, to varying states of cellular abnormalities that ultimately lead to cervical cancer. Lowgrade SIL (LSIL), like another cytological category - atypical squamous cells of undetermined significance (ASCUS), may progress to high-grade SIL (HSIL) and invasive cervical cancer or may regress to a normal state. Methods: In this work, we studied cervical canal microbiome in 165 HPVpositive and HPV-negative women of a reproductive age with ASCUS [HPV(+) n = 29; HPV(-) n = 11], LSIL [HPV(+) n = 32; HPV(-) n = 25], HSIL [HPV(+) n = 46], and the control group with negative for intraepithelial lesion malignancy (NILM) [HPV(-) n = 22]. Results and Discussion: HPV16 is the most prevalent HPV type. We have not found any differences between diversity in studied groups, but several genus [like Prevotella (p-value = 0.026), Gardnerella (p-value = 0.003), Fannyhessea (p-value = 0.024)] more often occurred in HSIL group compared by NILM or LSIL regardless of HPV. We have found statistically significant difference in occurrence or proportion of bacterial genus in studied groups. We also identified that increasing of the ratio of Lactobacillus iners or age of patient lead to higher chance to HSIL, while increasing of the ratio of Lactobacillus crispatus lead to higher chance to LSIL. Patients with a moderate dysbiosis equally often had either of three types of vaginal microbial communities (CST, Community State Type) with the prevalence of Lactobacillus crispatus (CST I), Lactobacillus gasseri (CST II), and Lactobacillus iners (CST III); whereas severe dysbiosis is linked with CST IV involving the microorganisms genera associated with bacterial vaginosis and aerobic vaginitis: Gardnerella, Fannyhessea, Dialister, Sneathia, Anaerococcus, Megasphaera, Prevotella, Finegoldia, Peptoniphilus, Porphyromonas, Parvimonas, and Streptococcus. [ABSTRACT FROM AUTHOR]

Programmed death-ligand 1 (PD-L1) is overexpressed in cervical carcinoma, hindering tumor destruction. The aim of this study was to assess PD-L1 expression by immunohistochemistry in cervical squamous cell carcinoma (SCC) and squamous intraepithelial lesions (SILs) from human immunodeficiency virus–positive (HIV+) and human immunodeficiency virus-negative (HIV-) patients. A total of 166 SCC and SIL samples of HIV+ and HIV- patients were included and analyzed for PD-L1 expression through tumor proportion score (TPS), and results were stratified in five TPS groups using SP263 antibody and, combined positive score (CPS) using 22C3 antibody. In cohort 1 (SP263 clone), all HIV+ patients were negative for intraepithelial lesion or malignancy (NILM), and low-grade squamous intraepithelial lesions (LSILs) scored < 1; and 87.5% of high-grade squamous intraepithelial lesions (HSILs) adjacent to SCC, 19% of HSILs non-adjacent to SCC, and 69% of SCCs scored ≥ 1 (15.4% scored 5). In HIV- patients, all NILM, LSILs, HSILs adjacent to SCC, and two HSILs non-adjacent to SCC scored < 1. SCC: 88.2% scored ≥ 1 and 5.9% scored 5. In cohort 2 (SP263 and 22C3 clones), 16.7% of HIV+ patients with SCC were positive with both clones, CPS ≥ 1 (22C3) or score 5 (≥ 50%) (SP263), showing no significant differences in positivity between both clones. These results indicate that a relatively low percentage of SCCs (16.7%; both in HIV+ and in HIV- patients) express PD-L1 (TPS ≥ 50% and CPS > 1), which may be due to some samples being archival material, sample characteristics, or use of different methodologies, highlighting the need for standardization of PD-L1 assessment in SCC of the cervix. The fact that PD-L1 is overexpressed in SILs of HIV+ patients suggests potential additional applications for immunotherapy in this disease. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To determine the association between apoptosis and the histopathological parameters that define squamous intraepithelial lesion of the uterine cervix. MATERIALS AND METHODS: Observational, cross-sectional, and relational study, by histopathological study of cervical biopsies with colposcopic suspicion of squamous intraepithelial lesion received at the Department of Pathology in the years 2018-2021. The following parameters were evaluated: apoptosis, koilocytes, number of mitoses, mitosis height, atypical mitosis, chromatin, basaloid atypia, nucleolus, and intraepithelial neutrophils. With the SPSS software, Pearson's Chi-square, Phi, Cramer's V and probability ratio for apoptosis were calculated with each of the parameters. Subsequently, the association and the measurement of the strength of association between apoptosis and the histopathological diagnosis of the studied cases were determined. RESULTS: A total of 624 glass slides were studied. A significant and relatively strong association was observed between apoptosis and bi-multinucleations, chromatin alterations, increased mitosis, and mitosis above the suprabasal layer, and a significant and moderate association between apoptosis and koilocytes, basaloid atypia, atypical mitosis, overlap and nuclear density. A significant and strong association was demonstrated between apoptosis and histopathological diagnosis of high-grade squamous intraepithelial lesion (Cramer's V: 701, p <.001) and a significant but weak and negative association with low-grade intraepithelial lesion (Phi: -.263, p <.001). CONCLUSIONS: Apoptosis is strongly associated with high-grade squamous intraepithelial lesion, and weakly and negatively associated with low-grade lesion. [ABSTRACT FROM AUTHOR]

Cervical cytology has been an integral part of cervical cancer screening since the mid‐20th century with the implementation of screening protocols utilising Pap testing. During that time, cervical cancer has gone from the leading cause of cancer deaths in women to not even appearing in the top 10 causes of US cancer deaths. However, despite its long and widespread use, cervical cytology remains a diagnostically challenging area in the practice of cytopathology. Of particular importance for diagnosticians is the accurate diagnosis of high‐grade squamous intraepithelial lesions (HSILs), given the significant risk of progression to invasive cervical cancer and the importance to patient management. Therefore, this review is presented in order to highlight the diagnostic features of HSIL, its various appearances, and important benign and neoplastic differential considerations with an emphasis on morphological clues that can aid in distinguishing between these different processes. [ABSTRACT FROM AUTHOR]

Abstract The lack of human‐derived in vitro models that recapitulate cervical pre‐cancerous lesions has been the bottleneck in researching human papillomavirus (HPV) infection‐associated pre‐cancerous lesions and cancers for a long time. Here, a long‐term 3D organoid culture protocol for high‐grade squamous intraepithelial lesions and cervical squamous cell carcinoma that stably recapitulates the two tissues of origin is described. Originating from human‐derived samples, a small biobank of cervical pre‐tumoroids and tumoroids that faithfully retains genomic and transcriptomic characteristics as well as the causative HPV genome is established. Cervical pre‐tumoroids and tumoroids show differential responses to common chemotherapeutic agents and grow differently as xenografts in mice. By coculture organoid models with peripheral blood immune cells (PBMCs) stimulated by HPV antigenic peptides, it is illustrated that both organoid models respond differently to immunized PBMCs, supporting organoids as reliable and powerful tools for studying virus‐specific T‐cell responses and screening therapeutic HPV vaccines. In this study, a model of cervical pre‐cancerous lesions containing HPV is established for the first time, overcoming the bottleneck of the current model of human cervical pre‐cancerous lesions. This study establishes an experimental platform and biobanks for in vitro mechanistic research, therapeutic vaccine screening, and personalized treatment for HPV‐related cervical diseases.

IntroductionHuman papilloma virus (HPV) is the most common sexually transmitted infection worldwide. Cervicovaginal microbiota plays an important role in HPV infection and is associated with the development of squamous intraepithelial lesions (SIL). The natural history of cervical cancer involves reversible changes in the cervical tissue from a normal state, in which no neoplastic changes are detected in the squamous epithelium, to varying states of cellular abnormalities that ultimately lead to cervical cancer. Low-grade SIL (LSIL), like another cytological category - atypical squamous cells of undetermined significance (ASCUS), may progress to high-grade SIL (HSIL) and invasive cervical cancer or may regress to a normal state.MethodsIn this work, we studied cervical canal microbiome in 165 HPV-positive and HPV-negative women of a reproductive age with ASCUS [HPV(+) n = 29; HPV(−) n = 11], LSIL [HPV(+) n = 32; HPV(−) n = 25], HSIL [HPV(+) n = 46], and the control group with negative for intraepithelial lesion malignancy (NILM) [HPV(−) n = 22].Results and DiscussionHPV16 is the most prevalent HPV type. We have not found any differences between diversity in studied groups, but several genus [like Prevotella (p-value = 0.026), Gardnerella (p-value = 0.003), Fannyhessea (p-value = 0.024)] more often occurred in HSIL group compared by NILM or LSIL regardless of HPV. We have found statistically significant difference in occurrence or proportion of bacterial genus in studied groups. We also identified that increasing of the ratio of Lactobacillus iners or age of patient lead to higher chance to HSIL, while increasing of the ratio of Lactobacillus crispatus lead to higher chance to LSIL. Patients with a moderate dysbiosis equally often had either of three types of vaginal microbial communities (CST, Community State Type) with the prevalence of Lactobacillus crispatus (CST I), Lactobacillus gasseri (CST II), and Lactobacillus iners (CST III); whereas severe dysbiosis is linked with CST IV involving the microorganisms genera associated with bacterial vaginosis and aerobic vaginitis: Gardnerella, Fannyhessea, Dialister, Sneathia, Anaerococcus, Megasphaera, Prevotella, Finegoldia, Peptoniphilus, Porphyromonas, Parvimonas, and Streptococcus.

Background: High-grade anal intraepithelial squamous lesion is significantly prevalent among men who have sex with men and are infected with the human immunodeficiency virus (HIV). This condition-the precursor to anal cancer-significantly increases the risk of developing it. Conversely, low-grade anal intraepithelial squamous typically follow a benign course and usually regress spontaneously., Materials and Methods: To describe a population of men who have sex with men living with HIV followed in a specialized anal cancer screening unit we conducted an observational, retrospective, and single-center study., Results: Ninety-four patients were analyzed, with a mean age of 39±9 years, and a 87% positivity rate for high-risk human papillomavirus (HR-HPV). At the initial visit, 47% presented with low-grade squamous intraepithelial lesions. The progression rate to high-grade squamous intraepithelial lesion was 37.2 per 100,000 patients/year. None of the patients developed anal cancer. Tobacco and alcohol consumption were associated with this progression., Discussion: In this series, longer duration of HIV infection, tobacco and alcohol use and the presence of HR-HPV were significantly associated with the occurrence of high-grade intraepithelial lesions. A lower risk of progression was seen in patients with higher education., Conclusion: In men who have sex with men living with HIV, the association of factors such as smoking, alcohol, the presence of HR-HPV and an increased burden of human papillomavirus disease makes these patients more susceptible to develop high-grade anal squamous lesions., (Copyright © 2024. Publicado por Elsevier España, S.L.U.)

Introduction: Dual immunostaining for p16/Ki67 is FDA-approved for use on liquid-based cervical cytology specimens; however, the utility of dual staining in anal cytology especially for ASCUS risk stratification is not well established., Methods: We investigated dual staining performance on anal cytology specimens and correlated with subsequent cytologic interpretation, high-risk HPV status, and anal biopsy results. Dual staining for p16/Ki-67 was performed on all liquid-based anal cytology specimens from December 2021 to June 2022 (n = 43)., Results: Three patients had high grade squamous intraepithelial lesion (HSIL/AIN2-3) on biopsy; dual staining was positive in all three cases. All HR-HPV negative cases were negative for dual staining. Among the 12 ASCUS samples with subsequent anal biopsy results all also had HR-HPV testing. Due to small sample size of cases with squamous intraepithelial lesion (SIL) diagnosed on biopsy, the sensitivity and positive predictive value was not calculated. However, the specificity and negative predictive value of p16/Ki-67 dual staining for SIL of any grade on biopsy were 1 (95% CI: 0.66-1) and 0.9 (95% CI: 0.65-0.97) respectively, whereas the specificity and negative predictive value of HR-HPV testing for SIL of any grade on biopsy were 0.44 (95% CI: 0.14-0.79) and 0.8 (95% CI: 0.41-0.96) respectively., Conclusion: Dual p16/Ki-67 staining indicates transforming HPV infection and could help serve as an ancillary test for risk stratification for atypical anal cytology specimens. Among ASCUS samples, dual staining was specific for SIL of any grade with a high negative predictive value and therefore could be useful in clinical practices with limited availability for follow-up care., (© 2024 Wiley Periodicals LLC.)

Background: High-grade anal intraepithelial squamous lesion is significantly prevalent among men who have sex with men and are infected with the human immunodeficiency virus (HIV). This condition-the precursor to anal cancer-significantly increases the risk of developing it. Conversely, low-grade anal intraepithelial squamous typically follow a benign course and usually regress spontaneously., Materials and Methods: To describe a population of men who have sex with men living with HIV followed in a specialized anal cancer screening unit we conducted an observational, retrospective, and single-center study., Results: Ninety-four patients were analyzed, with a mean age of 39±9 years, and a 87% positivity rate for high-risk human papillomavirus (HR-HPV). At the initial visit, 47% presented with low-grade squamous intraepithelial lesions. The progression rate to high-grade squamous intraepithelial lesion was 37.2 per 100,000 patients/year. None of the patients developed anal cancer. Tobacco and alcohol consumption were associated with this progression., Discussion: In this series, longer duration of HIV infection, tobacco and alcohol use and the presence of HR-HPV were significantly associated with the occurrence of high-grade intraepithelial lesions. A lower risk of progression was seen in patients with higher education., Conclusion: In men who have sex with men living with HIV, the association of factors such as smoking, alcohol, the presence of HR-HPV and an increased burden of human papillomavirus disease makes these patients more susceptible to develop high-grade anal squamous lesions., (Copyright © 2024. Publicado por Elsevier España, S.L.U.)

Cervical squamous intraepithelial lesions (SILs) may present a diagnostic challenge due to their morphological similarity to benign conditions and variability in interpretation, necessitating the exploration of objective biomarkers to aid in their identification and grading. This study evaluates the immunohistochemical markers Carbonic Anhydrase IX (CAIX) and ProEx-C to assess their diagnostic potential in cervical SILs. We retrospectively identified 56 SIL cases, including 20 low-grade SILs (LSIL) and 36 high-grade SILs (HSIL), alongside a control group of nine chronic cervicitis cases. Immunohistochemical staining was performed on formalin-fixed, paraffin-embedded tissue blocks, and the sensitivity and specificity of CAIX and ProEx-C were evaluated for detecting and grading SILs. CAIX exhibited a sensitivity of 64.3 % and a specificity of 100 % for detecting SILs. ProEx-C staining displayed a sensitivity of 66.1 % and a specificity of 100 % for detecting SILs. The combination of CAIX and ProEx-C staining increased the sensitivity for detecting SILs to 80.4 % without compromising the specificity. These markers alone or in combination were not found to be significant in distinguishing LSIL from HSIL. Both markers showed positivity in benign endocervical and squamous epithelium in high rates. In conclusion, CAIX and ProEx-C are valuable immunohistochemical markers for detecting SILs in cervical specimens, with high sensitivity and specificity. Further research is needed to elucidate their roles in cervical carcinogenesis and their relationship with HPV infection. • CAIX and ProEx-C show high sensitivity and specificity for detecting squamous intraepithelial lesions (SILs). • CAIX and ProEx-C do not reliably differentiate between LSILs and HSILs. • Benign cells are often positive for CAIX and ProEx-C, hence, correlating staining with cytomorphology remains essential. [ABSTRACT FROM AUTHOR]

Programmed death-ligand 1 (PD-L1) is overexpressed in cervical carcinoma, hindering tumor destruction. The aim of this study was to assess PD-L1 expression by immunohistochemistry in cervical squamous cell carcinoma (SCC) and squamous intraepithelial lesions (SILs) from human immunodeficiency virus-positive (HIV+) and human immunodeficiency virus-negative (HIV-) patients. A total of 166 SCC and SIL samples of HIV+ and HIV- patients were included and analyzed for PD-L1 expression through tumor proportion score (TPS), and results were stratified in five TPS groups using SP263 antibody and, combined positive score (CPS) using 22C3 antibody. In cohort 1 (SP263 clone), all HIV+ patients were negative for intraepithelial lesion or malignancy (NILM), and low-grade squamous intraepithelial lesions (LSILs) scored < 1; and 87.5% of high-grade squamous intraepithelial lesions (HSILs) adjacent to SCC, 19% of HSILs non-adjacent to SCC, and 69% of SCCs scored ≥ 1 (15.4% scored 5). In HIV- patients, all NILM, LSILs, HSILs adjacent to SCC, and two HSILs non-adjacent to SCC scored < 1. SCC: 88.2% scored ≥ 1 and 5.9% scored 5. In cohort 2 (SP263 and 22C3 clones), 16.7% of HIV+ patients with SCC were positive with both clones, CPS ≥ 1 (22C3) or score 5 (≥ 50%) (SP263), showing no significant differences in positivity between both clones. These results indicate that a relatively low percentage of SCCs (16.7%; both in HIV+ and in HIV- patients) express PD-L1 (TPS ≥ 50% and CPS > 1), which may be due to some samples being archival material, sample characteristics, or use of different methodologies, highlighting the need for standardization of PD-L1 assessment in SCC of the cervix. The fact that PD-L1 is overexpressed in SILs of HIV+ patients suggests potential additional applications for immunotherapy in this disease., (© 2023. The Author(s).)

Objective: In this study, we established a Sprague-Dawley rat model of vulvar squamous intraepithelial lesions and investigated the impact of focused ultrasound on the expression of hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF) and mutant type p53 (mtp53) in the vulvar skin of rats with low-grade squamous intraepithelial lesions (LSIL)., Materials and Methods: The vulvar skin of 60 rats was treated with dimethylbenzanthracene (DMBA) and mechanical irritation three times a week for 14 weeks. Rats with LSIL were randomly allocated into the experimental group or the control group. The experimental group was treated with focused ultrasound, while the control group received sham treatment., Results: After 14 weeks treatment of DMBA combined with mechanical irritation, LSIL were observed in 44 (73.33%) rats, and high-grade squamous intraepithelial lesions (HSIL) were observed in 14 (23.33%) rats. 90.91% (20/22) of rats showed normal pathology and 9.09% (2/22) of rats exhibited LSIL in the experimental group at four weeks after focused ultrasound treatment. 22.73% (5/22) of rats exhibited LSIL, 77.27% (17/22) of rats progressed to HSIL in the control group. Compared with the control-group rats, the levels of HIF-1α, VEGF and mtp53 were significantly decreased in experimental-group rats ( p  < 0.05)., Conclusions: These results indicate that DMBA combined with mechanical irritation can induce vulvar squamous intraepithelial lesion in SD rats. Focused ultrasound can treat LSIL safely and effectively, prevent the progression of vulvar lesions, and improve the microenvironment of vulvar tissues by decreasing the localized expression of HIF-1α, VEGF, and mtp53 in rats.