An in vitro model and the underlying pathways of sinonasal inverted papilloma development.

Recently, the specific association between Sinonasal inverted papilloma (SIP) and EGFR exon 20 mutations has been reported. To investigate the link between specific EGFR mutations and SIP development, we established organotypic raft culture system using nasal polyp-derived immortalized NP2 (iNP2) cells expressing EGFR exon 20 mutants or an exon 19 mutant, and SIP-derived iIP4 cells harboring P772_H773insPYNP mutation. In the raft culture, iIP4 cells showed the inverted growth pattern characteristic to SIP. Interestingly, iNP2 cells expressing EGFR exon 20 duplication mutants, S768_D770dup and N771_H773dup, but not of EGFR exon 19 mutant, E746_A750del, showed the inverted growth pattern. Enhanced activation of the PI3K/AKT signaling pathway was observed in iNP2_S768_D770dup and iIP4 cells, while increased MAPK signaling was found in iNP2_N771_H773dup. Increased cell migration and invasion were found in all cells carrying EGFR mutations when compared to iNP2 cells, and this effect was inhibited by either PI3K or MEK inhibitor. Notably, iNP2 cells expressing the N771_H773dup mutant showed the highest migration and invasion abilities. These results suggest that specific mutations in EGFR exon 20 play a crucial role in SIP development, partially though hyper-activation of the PI3K/AKT and MAPK signaling pathways. This study presents the first in vitro model for SIP development, which could facilitate further investigations into SIP pathogenesis and preclinical studies for new therapeutic agents.

Establishment and genetic characterization of cell lines derived from proliferating nasal polyps and sinonasal inverted papillomas.

To better understand the pathogenesis of nasal polyps (NPs) and sinonasal inverted papillomas (SIPs), we aimed to establish cell lines from fresh tissues of NPs and SIPs and characterize them. Primary cell cultures were obtained from two NP tissues (NP2 and NP3) and one SIP tissue (IP4). All the cells were polygonal in shape, expressed cytokeratin 14, and had normal diploid chromosome status. HPV58 DNA was detected in NP3. To obtain immortal primary cells, NP2 and IP4 cells were transduced with a combination of mutant CDK4, cyclinD1 and TERT. These cells were thereafter named NP2/K4DT and IP4/K4DT, respectively. HPV58-positive NP3 cells were transduced with TERT alone, the resulting cells named NP3/T. Phenotypic and genotypic identity of original tissues and derived cells was investigated. All the cell cultures with transgenes were confirmed to be derived from their parental cells and primary tumor tissues by analysis of short tandem repeats (STR) and maintained in vitro growth, genetic profiles and gene expression characteristics of the primary cells. These virtually immortalized cells, as well as the primary cells, have potential as in vitro models for studying the pathogenesis of NPs and SIPs and for preclinical study to develop new therapeutic agents.

Prevalence and association of Epstein-Barr virus infection with sinonasal inverted papilloma and sinonasal squamous cell carcinoma in the northeastern Thai population.

AIMS: Sinonasal inverted papillomas (SIP) and sinonasal squamous cell carcinomas (SNSCC) are sinonasal tumors with unclear etiology and pathogenesis. Epstein-Barr virus (EBV) has been detected in these tumors but information concerning their association is still limited. This study aimed to investigate the prevalence in, and association of EBV infection with SIP and SNSCC in northeastern Thailand. METHODS: DNA was extracted from 226 formalin-fixed, paraffin-embedded tissues including 80 nasal polyps (NP; the control group), 64 SIP and 82 SNSCC samples. Presence of EBV in these tissues was investigated using real-time PCR and their localization within tissues was confirmed using in situ hybridization (ISH). Characteristics of patients and the association of EBV prevalence with sinonasal tumors were analyzed. RESULTS: SIP and SNSCC were frequently found in people aged > 50 years and more often in males than in females (3:1 ratio). EBV infection was detected in 33.75, 64.06 and 37.80% of NP, SIP and SNSCC tissues, respectively, by real-time PCR. There was a statistically significant association between EBV infection and SIP (odds ratio [OR] = 3.52). This was not the case for SNSCC when compared to the NP group (OR = 1.83). Interestingly, EBV infection tended to be associated with inflammation and dysplasia in SIP. In SNSCC, EBV was mostly found in samples with undifferentiated or poorly differentiated cell types as well as in recurrent cases and lymph-node metastasis. Using ISH, EBV was detected only in infiltrating lymphocytes within the tumor stroma, not in the tumor epithelial cells. CONCLUSIONS: Infiltrating lymphocytes containing EBV in the tumor microenvironment might enhance tumorigenesis of SIP and SNSCC. The mechanism by which EBV promotes development of SIP and SNSCC needs to be elucidated in the future.

Chlamydia Trachomatis Infection in High-Risk Human Papillomavirus Based on Cervical Cytology Specimen.

OBJECTIVE: High-risk human papillomavirus (HR HPV) was associated with the development of cervical cancer. Asymptomatic Chlamydia trachomatis (C. trachomatis) infection is the most common bacterial, sexually-transmitted infection. This study aimed to investigate the association of C. trachomatis in positive HR HPV and the cytological results from liquid-based cytology (LBC). METHODS: 150 residual LBC specimens were collected; all of which had undergone cytology and HPV testing by Cobas. The samples were established as C. trachomatis using real-time PCR (RT-PCR) with Cryptic F/Cryptic R primers. RESULTS: Of 150 positive HPV findings, the most common (72.7%, 109/150) were the 12 other HR HPVs (viz., 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and 68). The cervical cytology of those positive HR HPVs were mostly negative (70.0%, 105/150).  The C. trachomatis infections in positive HR HPV were 16% (24/150) HPV. The analysis of the abnormal cytology revealed that 41.6% had C. trachomatis co-infection (C. trachomatis and HPV infection) viz., LSIL (20.8%), HSIL (12.5%), and ASC-US (8.3%). A comparison with positive HPV without C. trachomatis co-infection revealed that the highest prevalence was for LSIL, ASC-US, and HSIL (11.1%, 10.3%, and 6.4%, respectively). There was no difference between the abnormalities and negative cervical cytology with negative and positive C. trachomatis co-infection in HR HPV positive (p = 0.174). CONCLUSION: C. trachomatis infection was not significantly associated HR-HPV and abnormal cytology. This study confirms the increasing rate of C. trachomatis infection in asymptomatic women so routine screening for these infections has been suggested to (a) prevent complications such as the chronic pelvic pain associated with prolong infection and (b) reduce sexual transmission of the infection.

Modified Liquid-Based Cytology Technique for Immunocytochemistry in Effusion Specimen.

Objective: Immunocytochemistry (ICC) of serous effusion is an important tool for the diagnosis of benign and malignant cells. Our aim was to develop a modified liquid-based cytological technique for ICC (i.e., a modified LBC). Methods: Serous effusions of 110 cases were collected for cytological examination: 50 were negative for malignancy albeit benign mesothelium was found, and 60 were confirmed metastatic adenocarcinoma according to the modified LBC preparation. The latter were stained for EMA, Ber-EP4, Calretinin, and p63 then interpreted by both a cytotechnologist and a pathologist. A comparative analysis of the diagnostic results was conducted. Results: The results of the metastatic adenocarcinoma were 100% (60/60) positive for EMA and 91.7% (55/60) positive for Ber-Ep4 but negative for calretinin and p63. Cases negative for malignancy were 100% (50/50) positive for calretinin but negative for carcinoma markers. The difference between 'positive for metastatic adenocarcinoma' and 'negative for malignancy' in ICC was statistically significant (p < 0.001). Conclusion: The current study demonstrated that a panel marker, comprising EMA, Ber-EP4, and calretinin can be used for differentiating between cases of metastatic adenocarcinoma and benign mesothelium. The serous effusion specimen collected by the modified LBC technique is an effective preparation method for ICC.

Indocyanine green dye angiography as an adjunct to assess indeterminate burn wounds: A prospective, multicentered, triple-blinded study.

BACKGROUND: Clinical assessment of indeterminate burn wounds has been reported to yield poor accuracy, even when performed by burn experts. Indocyanine green (ICG) dye angiography has been found to be highly accurate in assessing burn depth, but there is still limited evidence of its use in indeterminate burn wounds. This study aims to compare the accuracy of ICG angiography to that of clinical assessment in assessing indeterminate burn wounds. METHODS: This is a prospective, multicentered, triple-blinded, experimental study. Participants were stable patients, admitted to the hospital with burn wounds of indeterminate depth. The burn wounds were clinically assessed by an attending plastic surgeon. ICG angiography was performed and evaluated by another surgeon. Tissue biopsies were obtained and sent for histological study to be assessed as the gold standard. RESULTS: In the 30 burn sites that were assessed, the accuracy of ICG angiography was 100.0%, compared with 50.0% for clinical assessment (p < 0.001). Clinical assessment yielded a sensitivity of 33.3% and specificity of 66.7%, while ICG angiography yielded both a sensitivity and specificity of 100.0%. Therefore, the number needed to treat for using ICG angiography in indeterminate burn wounds was two. CONCLUSION: Indocyanine green angiography yields a significantly higher accuracy than clinical assessment in indeterminate burn wounds. This intervention can, thus, be a useful tool to aid clinical judgment. TRIAL REGISTRATION: Thai Clinical Trials Registry, number TCTR20170821001. LEVEL OF EVIDENCE: Diagnostic test, level I.

Comparative Analysis of Modified Liquid-Based Cytology and CytoRich Red Preparation in Assessment of Serous Effusion for Cancer Diagnosis

Objective: We aimed to compare the cytomorphological diagnosis in serous effusion and quality of background between modified liquid-based cytology (modified-LBC) and CytoRich Red (CRR) preservative. Methods: We used an experimental study design: 110 fresh serous effusions were received from 50 cases negative for malignant effusions and 60 cases positive for malignant effusions. All fresh serous effusions were processed using both the CRR solution and the modified-LBC preparation. Blind sample slides were interpreted for cytomorphological diagnosis and the quality of background by 2 cytotechnologists. Result: All cases had the same diagnosis irrespective of the method. There was no statistically significant difference in the cytological diagnosis between the CRR and modified-LBC preparations (p>0.999). The quality of the background smear for the CRR preparation was clean (54%), moderate in 42%, and poor in 4%. By comparison, the modified-LBC preparation was clean in 46%, moderate in 47%, and poor in 7%. The difference between the quality of background smears between the two methods was not statistically significant (p= 0.527). Conclusion: There was no statistically significant difference in the diagnosis or quality of background between CRR and modified-LBC preparations. The serous effusion specimen prepared by modified-LBC solution was less expensive than CRR. The modified-LBC could be an alternative preparation when commercial preparations are too expensive.

Malignant mesothelioma of the pericardium: a report of two different presentations.

Malignant mesothelioma of the pericardium is a rare and fatal condition that clinicians should be aware of due to its variability of clinical manifestation. The diagnosis may be delayed as a result of delayed treatment. Here, we report two cases of malignant pericardial mesothelioma with two different clinical aspects: cardiac tamponade and mimic tuberculous pericarditis. Both patients: may have indirect exposure to asbestos. Despite chemotherapy, both patients died at 2 weeks and 3 months after the diagnosis. Malignant mesothelioma of the pericardium is fatal, has a variety of presentation, and may not be related to asbestosis exposure.