MicroRNA-21 (miR-21) is recognized as an oncomir and shows up-regulation in many types of human malignancy. The aim of this study was to investigate the association of miR-21 expression associated with HPV infection in normal and abnormal cervical tissues. Cervical tissue samples with different cytological or histopathological grades were investigated for HPV by PCR and for miR-21 and programmed cell death, protein 4 (PDCD4) expression using quantitative real-time PCR (qRT-PCR). Laser capture microdissection (LCM) of stromal and epithelial tissues and in situ hybridization (ISH) using locked nucleic acid (LNA) probes were performed on a subset of fixed specimens. Cell line experiments were conducted on fibroblasts stimulated in culture media from HeLa cells, which were then assessed for miR-21, PDCD4, IL-6 and α-SMA expression by qRT-PCR. Twenty normal cervical cell, 12 cervicitis, 14 cervical intraepithelial neoplastic I (CIN I), 22 CIN II-III and 43 cervical squamous cell carcinoma (SCC) specimens were investigated. miR-21 levels were significantly lower in normal than in abnormal tissues. The expression of miR-21 in HPV negative normal cytology was significantly lower than in HPV positive samples in abnormal tissue and SCC. The miR-21 expression was significantly higher in HPV negative cervicitis than HPV negative normal cells. LCM and ISH data showed that miR-21 is primarily expressed in the tumor-associated stromal cell microenvironment. Fibroblasts treated with HeLa cell culture media showed up-regulated expression of miR-21, which correlated with increased expression of α-SMA and IL-6 and with down-regulation of PDCD4. These results demonstrate that miR-21 is associated with HPV infection and involved in cervical lesions as well as cervicitis and its up-regulation in tumor-stroma might be involved in the inflammation process and cervical cancer progression.

HPV infected cervical cells secrete mediators that are gradually changed and have influence on infiltrating M2 phenotypic monocytes in cervical lesions. However, profiles of circulating immune cells in women with cervical lesions and M2 phenotypic monocyte activity in HPV infected cervical lesions are limited. This study aimed to investigate circulating monocyte populations correlated with M2 phenotype density and its activity in HPV infected cervical lesions. HPV DNA was investigated in cervical tissues using PCR. High risk HPV E6/E7 mRNA was detected using in situ hybridization. CD163 immunohistochemical staining was performed for M2 macrophage. CD163 and Arg1 mRNA expression were detected using real-time PCR. Circulating monocyte subpopulations were analyzed using flow cytometry. CD163 and Arg1 mRNA expression were increased according to cervical lesion severity and corresponding with density of M2 macrophage in HSIL and SCC in stroma and peri-tumoral areas. Additionally, the relationship between M2 macrophage infiltration and high risk HPV E6/E7 mRNA expression was found and corresponded with cervical lesion severity. Circulating CD14CD16 and CD14CD163 monocytes were elevated in No-SIL and cervical lesions. Interestingly, CD14CD64 monocyte was greatly elevated in HSIL and SCC, whereas intracellular IL-10 monocytes were not significantly different between cervical lesions. The correlation between increasing ratio of circulating CD64/CD163 monocyte and density of infiltrating CD163 monocytes was associated with severity of HPV infected cervical lesions. The elevated circulating CD64/CD163 monocyte ratio correlates to severity of HPV infected cervical lesions and might be a prognostic marker in cervical cancer progression.

BackgroundCervical squamous cell carcinoma (CSCC) is a major cause of female mortality worldwide. This study has examined epidermal growth factor receptor (EGFR) pathway markers that represent actionable pharmacological targets.MethodsHPV16 positive CSCCs (n = 105 patients) from Madhya Pradesh, India were screened for KRAS and PIK3CA mutations by PNA-clamp real-time PCR. Immunohistochemistry (IHC) was performed for EGFR, PIK3CA, PTEN, phospho-AKT, phospho-mTOR and phospho-44/42 MAPK (ERK1/2).ResultsKRAS mutations were detected in 0/91 (0%) and PIK3CA mutations in 19/95 (20.0%) informative specimens: exon 9, E542 (n = 3) and E545 (n = 15); exon 20, H1047R (n = 1). PIK3CA mutation detection was associated with older mean patient age [48.2 vs. 56.6 years (P = 0.007)] and with post-menopausal age: 5/45 (11.1%) patients <50 years vs. 14/50 (28.0%) patients ≥50 years (P = 0.045; OR = 3.11). EGFR expression was present in 60/101 (59.4%) CSCCs and was associated with PIK3CA mutation detection (P < 0.05) but not age (P > 0.05). EGFR and phospho-AKT staining showed associations with tumor grade and/or lymph node status (P < 0.05). Significant associations were not found for the other study markers (P > 0.05).ConclusionThese data show that PIK3CA mutation acquisition is related to patient age and EGFR expression. The absence of KRAS mutations supports the potential of anti-EGFR therapies for CSCC treatment. The relatively high PIK3CA mutation rates indicate that PI3K may be a therapeutic target for a significant subset of CSCC patients. Qualitatively distinct IHC staining profiles for the marker panel were noted patient to patient; however, across patients, consistent linear relationships between up- and downstream pathway markers were not observed. Evaluation of the expression status of potential cancer pathway targets may be of value in addition to molecular profiling for choosing among therapeutic options.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-015-0611-0) contains supplementary material, which is available to authorized users.

Human papillomavirus (HPV) infection is associated with several genetic alterations including oncogene amplification, leading to increased aggression of tumors. Recently, a relationship between HPV infection and oncogene amplification has been reported, but this finding remains controversial. This study therefore investigated relationships between HPV infection and amplification of genes in the epidermal growth factor receptor (EGFR) signaling cascade in oral squamous cell carcinoma (OSCC). Extracted DNA from 142 formalin-fixed paraffin-embedded (FFPE) OSCC tissues was performed to investigate the copy number of EGFR, KRAS, c-myc and cyclin D1 genes using real-time polymerase chain reaction (RT-PCR) and compared with calibrators. A tissue microarray of OSCC tissues was used for detection of c-Myc expression and HPV infection by immunohistochemistry and HPV E6/E7 RNA in situ hybridization, respectively. HPV infection was also investigated using PCR and RT-PCR. Of the 142 OSCC samples, 81 (57%) were HPV-infected cases. The most frequently amplified gene was c-myc (55.6%), followed by cyclin D1 (26.1%), EGFR (23.9%) and KRAS (19.7%). Amplification of c-myc was significantly associated with levels of its protein product. EGFR amplification was also significantly associated with amplification of genes in the signaling cascade: KRAS (50.0%), c-myc (34.2%) and cyclin D1 (46.0%). Interestingly, HPV infection was significantly associated with amplification of both EGFR (76.5%) and cyclin D1 (73.0%). Only cyclin D1 amplification was significantly associated with severity of OSCC histopathology. HPV infection may play an important synergistic role in amplification of genes in the EGFR signaling cascade, leading to increased aggression in oral malignancies.

Objectives Cutibacterium acnes is one of the common multifactorial causes that play an important role in the pathophysiology of acne vulgaris. We aimed to develop novel antimicrobial peptides for reduction of the hypercolonization. Methods Six cationic peptides were derived by de novo designation. The antimicrobial and cytotoxic activities of peptides were investigated. The peptide conformation was determined by circular dichroism spectrometry. The antimicrobial effects of peptides were evaluated using scanning electron microscopy (SEM), transmission electron microscopy (TEM) and DNA-binding ability assay. Results Among designed peptides, WSKK11 and WSRR11 were effective antimicrobials against C. acnes at MICs of 128 and 64 mg/L, respectively. The MICs of WSKK11 against Staphylococcus epidermidis, Staphylococcus aureus and Candida albicans were 8, 8 and 32 mg/L, while those of WSRR11 were 64, 32 and 32 mg/L, respectively. WSKK11 and WSRR11 were less toxic to human erythrocytes (&lt;2%) and not toxic to macrophages, keratinocytes and fibroblasts up to 512 mg/L. WSKK11 and WSRR11 mostly revealed the conformation of the undefined or random coil structures under mimicked environmental conditions. The peptides affected cell surfaces and cell membranes of C. acnes as well as possibly translocating through the cell membrane, observed by a combination of SEM and TEM, respectively. WSKK11 and WSRR11 had the ability to bind bacterial DNA. Conclusions The two novel antimicrobial peptides WSKK11 and WSRR11 are members of a new class of antimicrobial agents that could deal with acne problems. Therefore, the antimicrobial peptides may be promising novel active agents for dermatological, beauty and cosmeceutical applications.

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The current practice of determining histologic grade with a single molecular biomarker can facilitate differential diagnosis but cannot predict the risk of lesion progression. Cancer is caused by complex mechanisms, and no single biomarker can both make accurate diagnoses and predict progression risk. Modelling using multiple biomarkers can be used to derive scores for risk prediction. Mathematical models (MMs) may be capable of making predictions from biomarker data. Therefore, this study aimed to develop MM–based scores for predicting the risk of precancerous cervical lesion progression and identifying precancerous lesions in patients in northern Thailand by evaluating the expression of multiple biomarkers. The MMs (Models 1–5) were developed in the test sample set based on patient age range (five categories) and biomarker levels (cortactin, p16INK4A, and Ki–67 by immunohistochemistry [IHC], and HPV E6/E7 ribonucleic acid (RNA) by in situ hybridization [ISH]). The risk scores for the prediction of cervical lesion progression (“risk biomolecules”) ranged from 2.56–2.60 in the normal and low–grade squamous intraepithelial lesion (LSIL) cases and from 3.54–3.62 in cases where precancerous lesions were predicted to progress. In Model 4, 23/86 (26.7%) normal and LSIL cases had biomolecule levels that suggested a risk of progression, while 5/86 (5.8%) cases were identified as precancerous lesions. Additionally, histologic grading with a single molecular biomarker did not identify 23 cases with risk, preventing close patient monitoring. These results suggest that biomarker level–based risk scores are useful for predicting the risk of cervical lesion progression and identifying precancerous lesion development. This multiple biomarker–based strategy may ultimately have utility for predicting cancer progression in other contexts.

Background The incidence of oral cancers associated with human papillomavirus (HPV) has been increasing in recent years. Therefore, it is necessary to elucidate HPV prevalence in oral cells and exposure to risk factors in various age groups. Methods Oral rinse samples from healthy individuals in northern Thailand were investigated for HPV prevalence and genotyped using the polymerase chain reaction (GP5+/6+ primers) and DNA sequencing of the PCR products. Results Samples were collected from 594 participants between 4 and 60 years of age. HPV was detected in 3.7% of samples. The prevalence of HPV‐positive cases was 8.6% in the 31‐50 age group. HPV prevalence increased with age and was the highest (9.2%) in the 41‐50 age group, but decreased (to 3%) in the 51‐60 age group. Risk factors significantly associated with HPV‐positive cases included alcohol consumption, coffee drinking, sexual activity, and having children. HPV 16 and 18 were common genotypes, especially in the 31‐50 age group, and were associated with having sexual activity (odds ratio 19.0 [95% CI: 2.5‐142.5]). At follow‐up of some individuals in the 4‐10 age group, a 9‐year‐old child was found to be positive for HPV18. Conclusions These results suggest that HPV can be acquired at a young age and the prevalence peaks in the middle age class among healthy individuals in northern Thailand, especially in the 31‐50 age group.