Conditional Cooperativity in RAS Assembly Pathways on Nanodiscs and Altered GTPase Cycling.

Self-assemblies (i.e., nanoclusters) of the RAS GTPase on the membrane act as scaffolds that activate downstream RAF kinases and drive MAPK signaling for cell proliferation and tumorigenesis. However, the mechanistic details of nanoclustering remain largely unknown. Here, size-tunable nanodisc platforms and paramagnetic relaxation enhancement (PRE) analyses revealed the structural basis of the cooperative assembly processes of fully processed KRAS, mutated in a quarter of human cancers. The cooperativity is modulated by the mutation and nucleotide states of KRAS and the lipid composition of the membrane. Notably, the oncogenic mutants assemble in nonsequential pathways with two mutually cooperative 'α/α' and 'α/ß' interfaces, while α/α dimerization of wild-type KRAS promotes the secondary α/ß interaction sequentially. Mutation-based interface engineering was used to selectively trap the oligomeric intermediates of KRAS and probe their favorable interface interactions. Transiently exposed interfaces were available for the assembly. Real-time NMR demonstrated that higher-order oligomers retain higher numbers of active GTP-bound protomers in KRAS GTPase cycling. These data provide a deeper understanding of the nanocluster-enhanced signaling in response to the environment. Furthermore, our methodology is applicable to assemblies of many other membrane GTPases and lipid nanoparticle-based formulations of stable protein oligomers with enhanced cooperativity.

Inhibition of protein-protein interactions using biodegradable depsipeptide nanoassemblies.

Although peptides notoriously have poor intrinsic pharmacokinetic properties, it is well-known that nanostructures with excellent pharmacokinetic properties can be designed. Noticing that peptide inhibitors are generally nonpolar, here, we consolidate the peptide inhibitor targeting intracellular protein-protein interactions (PPIs) as an integral part of biodegradable self-assembled depsipeptide nanostructures (SdPNs). Because the peptide inhibitor has the dual role of PPI inhibition and self-assembly in this design, problems associated with the poor pharmacokinetics of peptides and encapsulation/entrapment processes can be overcome. Optimized SdPNs displayed better tumor targeting and PPI inhibition properties than the comparable small molecule inhibitor in vivo. Kinetics of PPI inhibition for SdPNs were gradual and controllable in contrast to the rapid inhibition kinetics of the small molecule. Because SdPN is modular, any appropriate peptide inhibitor can be incorporated into the platform without concern for the poor pharmacokinetic properties of the peptide.

Clinical effects of bacteremia in sepsis patients with community-acquired pneumonia.

BACKGROUND: Data regarding the clinical effects of bacteremia on severe community-acquired pneumonia (CAP) are limited. Thus, we investigated clinical characteristics and outcomes of severe CAP patients with bacteremia compared with those of subjects without bacteremia. In addition, we evaluated clinical factors associated with bacteremia at the time of sepsis awareness. METHODS: We enrolled sepsis patients diagnosed with CAP at emergency departments (EDs) from an ongoing nationwide multicenter observational registry, the Korean Sepsis Alliance, between September 2019 and December 2020. For evaluation of clinical factors associated with bacteremia, we divided eligible patients into bacteremia and non-bacteremia groups, and logistic regression analysis was performed using the clinical characteristics at the time of sepsis awareness. RESULT: During the study period, 1,510 (47.9%) sepsis patients were caused by CAP, and bacteremia was identified in 212 (14.0%) patients. Septic shock occurred more frequently in the bacteremia group than in the non-bacteremia group (27.4% vs. 14.8%; p < 0.001). In multivariable analysis, hematologic malignancies and septic shock were associated with an increased risk of bacteremia. However, chronic lung disease was associated with a decreased risk of bacteremia. Hospital mortality was significantly higher in the bacteremia group than in the non-bacteremia group (27.3% vs. 40.6%, p < 0.001). The most prevalent pathogen in blood culture was Klebsiella pneumoniae followed by Escherichia coli in gram-negative pathogens. CONCLUSION: The incidence of bacteremia in severe CAP was low at 14.0%, but the occurrence of bacteremia was associated with increased hospital mortality. In severe CAP, hematologic malignancies and septic shock were associated with an increased risk of bacteremia.

mRNA-HPV vaccine encoding E6 and E7 improves therapeutic potential for HPV-mediated cancers via subcutaneous immunization.

The E6 and E7 proteins of specific subtypes of human papillomavirus (HPV), including HPV 16 and 18, are highly associated with cervical cancer as they modulate cell cycle regulation. The aim of this study was to investigate the potential antitumor effects of a messenger RNA-HPV therapeutic vaccine (mHTV) containing nononcogenic E6 and E7 proteins. To achieve this, C57BL/6j mice were injected with the vaccine via both intramuscular and subcutaneous routes, and the resulting effects were evaluated. mHTV immunization markedly induced robust T cell-mediated immune responses and significantly suppressed tumor growth in both subcutaneous and orthotopic tumor-implanted mouse model, with a significant infiltration of immune cells into tumor tissues. Tumor retransplantation at day 62 postprimary vaccination completely halted progression in all mHTV-treated mice. Furthermore, tumor expansion was significantly reduced upon TC-1 transplantation 160 days after the last immunization. Immunization of rhesus monkeys with mHTV elicited promising immune responses. The immunogenicity of mHTV in nonhuman primates provides strong evidence for clinical application against HPV-related cancers in humans. All data suggest that mHTV can be used as both a therapeutic and prophylactic vaccine.

The deubiquitinase UCHL3 mediates p300-dependent chemokine signaling in alveolar type II cells to promote pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic, fatal, fibrotic, interstitial lung disease of unknown cause. Despite extensive studies, the underlying mechanisms of IPF development remain unknown. Here, we found that p300 was upregulated in multiple epithelial cells in lung samples from patients with IPF and mouse models of lung fibrosis. Lung fibrosis was significantly diminished by the alveolar type II (ATII) cell-specific deletion of the p300 gene. Moreover, we found that ubiquitin C-terminal hydrolase L3 (UCHL3)-mediated deubiquitination of p300 led to the transcriptional activation of the chemokines Ccl2, Ccl7, and Ccl12 through the cooperative action of p300 and C/EBPß, which consequently promoted M2 macrophage polarization. Selective blockade of p300 activity in ATII cells resulted in the reprogramming of M2 macrophages into antifibrotic macrophages. These findings demonstrate a pivotal role for p300 in the development of lung fibrosis and suggest that p300 could serve as a promising target for IPF treatment.

Seasonal Dynamics of Bacterial Community Structure in Diesel Oil-Contaminated Soil Cultivated with Tall Fescue (Festuca arundinacea).

The objective of this study was to explore the seasonal characteristics of rhizoremediation and the bacterial community structure over the course of a year in soil contaminated with diesel oil. The soil was contaminated with diesel oil at a total petroleum hydrocarbon (TPH) concentration of 30,000 mg-TPH·kg-soil-1. Tall fescue seedlings were planted in the contaminated soil and rhizoremediation performance was monitored for 317 days. The TPH concentration gradually declined, reaching 75.6% after day 61. However, the TPH removability decreased by up to 30% after re-contamination in the fall and winter. The bacterial community structure exhibited distinct seasonal dynamics. Genus Pseudomonas significantly increased up to 55.7% in the winter, while the genera Immundisolibacter and Lysobacter, well-known petroleum hydrocarbon (PH)-degrading bacteria, were found to be positively linked to the TPH removal rate. Consequently, knowledge of this seasonal variation in rhizoremediation performance and the bacterial community structure is useful for the improvement of rhizoremediation in PH-contaminated environments.

A Novel Regulatory Role of Activated Leukocyte Cell-Adhesion Molecule in the Pathogenesis of Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive lung disease characterized by fibroproliferative matrix molecule accumulation, collagen deposition, and apoptosis. Activated leukocyte cell-adhesion molecule (ALCAM; CD166) is a cell-adhesion molecule that has been implicated in adhesive and migratory attribution, including leukocyte homing and trafficking and cancer metastasis. We investigated the role of ALCAM on pulmonary fibrosis development in murine models. Thus, a bleomycin-induced pulmonary fibrosis model was established with wild-type and ALCAM-/- mice. Pulmonary fibrosis was also induced in transforming growth factor-ß1 (TGF-ß1)-transgenic mice that conditionally overexpress TGF-ß1 upon doxycycline administration. In both models, observed reduced ALCAM levels in lung tissue and BAL fluid in pulmonary fibrosis-induced wild-type mice compared with control mice. We also observed reduced ALCAM expression in the lung tissue of patients with pulmonary fibrosis compared with normal lung tissue. ALCAM-/- mice showed an exacerbated lung fibrosis response compared with wild-type mice, and this was accompanied by increased cell death. Further investigation for identification of the signaling pathway revealed that PI3K and ERK signaling pathways are involved in bleomycin-induced fibrosis. Collectively, these results highlight that ALCAM plays a protective role in the pathogenesis of pulmonary fibrosis that inhibits epithelial cell apoptosis through the PI3K-Akt signaling pathway. Our findings demonstrate the potential of ALCAM as a therapeutic target for IPF and may aid the development of new strategies for the management and treatment of patients with IPF.

Club cell-specific role of programmed cell death 5 in pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) causes progressive fibrosis and worsening pulmonary function. Prognosis is poor and no effective therapies exist. We show that programmed cell death 5 (PDCD5) expression is increased in the lungs of patients with IPF and in mouse models of lung fibrosis. Lung fibrosis is significantly diminished by club cell-specific deletion of Pdcd5 gene. PDCD5 mediates ß-catenin/Smad3 complex formation, promoting TGF-ß-induced transcriptional activation of matricellular genes. Club cell Pdcd5 knockdown reduces matricellular protein secretion, inhibiting fibroblast proliferation and collagen synthesis. Here, we demonstrate the club cell-specific role of PDCD5 as a mediator of lung fibrosis and potential therapeutic target for IPF.

Prognostic value of tumor budding in gallbladder cancer: application of the International Tumor Budding Consensus Conference scoring system.

Tumor budding (TB), a histopathological manifestation of epithelial-mesenchymal transition, is an important step in cancer invasion and metastasis development. TB has been considered a strong prognostic indicator in colorectal cancer. The International Tumor Budding Consensus Conference (ITBCC) scoring system is the standardized method used for patient outcome prediction in several human tumors. We investigated the clinicopathological implications and applicability of TB measured using the ITBCC scoring system in gallbladder cancer (GBC). The TB grades assigned to the 78 GBC patients were as follows: Bd1 (low TB), 41 (52.6%) patients; Bd2 (intermediate TB), 22 (28.2%) patients; and Bd3 (high TB), 15 (19.2%) patients. A higher TB grade correlated with a poorer histological differentiation (P < 0.000), higher pT category (P < 0.000), the involvement of surgical resection margin (P = 0.005), presence of nodal metastasis (P < 0.000), lymphatic and venous invasion (P < 0.000), and perineural invasion (P = 0.004). Univariate Cox regression analysis revealed that a poor histological grade, high pT category, lymphatic invasion, perineural invasion, and intermediate to high TB grades were associated with worse 5-year overall survival and disease-free survival. TB was not significantly associated with death or recurrence risk in multivariate Cox analysis. The interobserver agreement of TB grading was substantial. This study is the first to apply the ITBCC scoring system and suggest the prognostic value of TB in GBC.

Aucuparin Suppresses Bleomycin-Induced Pulmonary Fibrosis Via Anti-Inflammatory Activity.

Idiopathic pulmonary fibrosis (IPF) is a lung disease that results in scarring of the lungs for an unknown reason. Although many studies have been conducted on IPF, precise mechanisms and treatments have not yet been identified. In this study, we found that aucuparin, a natural product isolated from Sorbus aucuparia, inhibited pulmonary fibrosis in a bleomycin (BLM)-induced lung fibrosis mouse model. In the lung samples of mice treated with aucuparin, the gene expression of inflammation and macrophage activation markers was reduced compared to those treated with BLM alone. Moreover, aucuparin decreased the expression of profibrotic marker genes and increased the expression of antifibrotic marker genes. Finally, we observed that aucuparin significantly suppressed transforming growth factor-ß-induced activation of inflammatory cytokine production and collagen synthesis from macrophages and fibroblasts, respectively. Taken together, these data demonstrate that aucuparin inhibits lung fibrosis via its anti-inflammatory action and support its potential to be a therapeutic drug for IPF treatment.

Interobserver diagnostic reproducibility in advanced-stage endometrial carcinoma.

BACKGROUND: The accurate pathologic diagnosis and subtyping of high-grade endometrial carcinoma are often problematic, due to its atypical and overlapping histopathological features. METHODS: Three pathologists reviewed 21 surgically resected cases of advancedstage endometrial carcinoma. The primary diagnosis was based only on hematoxylin and eosin stained slides. When a discrepancy arose, a secondary diagnosis was made by additional review of immunohistochemical (IHC) stains. Finally, three pathologists discussed all cases and rendered a consensus diagnosis. RESULTS: The primary diagnoses were identical in 13/21 cases (62%). The secondary diagnosis based on the addition of IHC results was concordant in four of eight discrepant cases. Among four cases with discrepancies occurring in this step, two cases subsequently reached a consensus diagnosis after a thorough discussion between three reviewers. Next-generation sequencing (NGS) study was performed in two cases in which it was difficult to distinguish between serous carcinoma and endometrioid carcinoma. Based on the sequencing results, a final diagnosis of serous carcinoma was rendered. The overall kappa for concordance between the original and consensus diagnosis was 0.566 (moderate agreement). CONCLUSIONS: We investigated stepwise changes in interobserver diagnostic reproducibility in advanced-stage endometrial carcinoma. We demonstrated the utility of IHC and NGS study results in the histopathological diagnosis of advanced-stage endometrial carcinoma.

Plumbagin Suppresses Pulmonary Fibrosis via Inhibition of p300 Histone Acetyltransferase Activity.

Idiopathic pulmonary fibrosis (IPF) is a chronic fibrosing interstitial lung disease with a poor prognosis similar to that of malignancy. The causes of IPF are not clearly known, and there is no effective therapy to date. In this study, the natural compound plumbagin, which was isolated from Plumbago rosea root extract, was screened for p300 inhibitory activity. Plumbagin specifically inhibited the activity of p300 toward histone acetyltransferases. Plumbagin treatment significantly suppressed transforming growth factor-ß-induced profibrotic target-gene expression and proliferation of fibroblast cell lines. Moreover, plumbagin significantly inhibited bleomycin-induced pulmonary fibrosis in mice. Taken together, these data demonstrate the inhibitory effects of plumbagin on lung fibrosis and its promise as a therapeutic agent for IPF.

Multiparametric MR Is a Valuable Modality for Evaluating Disease Severity of Nonalcoholic Fatty Liver Disease.

INTRODUCTION: Because nonalcoholic fatty liver disease (NAFLD) is becoming a leading cause of chronic liver disease, noninvasive evaluations of its severity are immediately needed. This prospective cross-sectional study evaluated the effectiveness of noninvasive assessments of hepatic steatosis, fibrosis, and steatohepatitis. METHODS: Patients underwent laboratory tests, liver biopsy, transient elastography, and MRI. Multiparametric MR was used to measure MRI proton density fat fraction, MR spectroscopy, T1 mapping, and MR elastography (MRE). RESULTS: We enrolled 130 patients between October 2016 and July 2019. For the diagnosis of moderate-to-severe steatosis (grade ≥ 2), the area under the receiver operating characteristic curve (AUROC) was lower in controlled attenuation parameter (0.69; 95% confidence interval [CI], 0.60-0.76) than MRI proton density fat fraction (0.82; 95% CI, 0.75-0.89; P = 0.008) and MR spectroscopy (0.83; 95% CI, 0.75-0.89; P = 0.006). For the diagnosis of advanced fibrosis (stage ≥ 3), the AUROC of MRE (0.89; 95% CI, 0.83-0.94) was superior compared with those of the Fibrosis-4 index (0.77; 95% CI, 0.69-0.84; P = 0.010), NAFLD fibrosis score (0.81; 95% CI, 0.73-0.87; P = 0.043), and transient elastography (0.82; 95% CI, 0.74-0.88; P = 0.062). For detecting advanced fibrosis or nonalcoholic steatohepatitis, the AUROC of MRE (0.86; 95% CI, 0.79-0.91) was higher than that of TE (0.76; 95% CI, 0.68-0.83) with statistical significance (P = 0.018). DISCUSSION: Multiparametric MR accurately identified a severe form of NAFLD. Multiparametric MR can be a valuable noninvasive method for evaluating the severity of NAFLD.

Multiparametric MR Index for the Diagnosis of Non-Alcoholic Steatohepatitis in Patients with Non-Alcoholic Fatty Liver Disease.

Non-alcoholic steatohepatitis (NASH) is a complex disease consisting of various components including steatosis, lobular inflammation, and ballooning degeneration, with or without fibrosis. Therefore, it is difficult to diagnose NASH with only one imaging modality. This study was aimed to evaluate the feasibility of magnetic resonance imaging (MRI) for predicting NASH and to develop a non-invasive multiparametric MR index for the detection of NASH in non-alcoholic fatty liver disease (NAFLD) patients. This prospective study included 47 NAFLD patients who were scheduled to undergo or underwent ultrasound-guided liver biopsy within 2 months. Biopsy specimens were graded as NASH or non-NASH. All patients underwent non-enhanced MRI including MR spectroscopy (MRS), MR elastography (MRE), and T1 mapping. Diagnostic performances of MRS, MRE, and T1 mapping for grading steatosis, activity, and fibrosis were evaluated. A multiparametric MR index combining fat fraction (FF), liver stiffness (LS) value, and T1 relaxation time was developed using linear regression analysis. Receiver operating characteristic (ROC) curve analysis was performed to evaluate the diagnostic performance of the newly devised MR index. Twenty NASH patients and 27 non-NASH patients were included. Using MRS, MRE, and T1 mapping, the mean areas under the curve (AUCs) for grading steatosis, fibrosis, and activity were 0.870, 0.951, and 0.664, respectively. The multiparametric MR index was determined as 0.037 × FF (%) + 1.4 × LS value (kPa) + 0.004 × T1 relaxation time (msec) -3.819. ROC curve analysis of the MR index revealed an AUC of 0.883. The cut-off value of 6 had a sensitivity of 80.0% and specificity of 85.2%. The multiparametric MR index combining FF, LS value, and T1 relaxation time showed high diagnostic performance for detecting NASH in NAFLD patients.

The Association between Low Vitamin D Status and Autoimmune Thyroid Disease in Korean Premenopausal Women: The 6th Korea National Health and Nutrition Examination Survey, 2013-2014.

BACKGROUND: This study aimed to analyze the association of low vitamin D status with thyroid autoimmunity and dysfunction in the Korean population according to sex and menopausal status in women. METHODS: This study was based on the data acquired from the 6th Korea National Health and Nutrition Examination Survey. We enrolled 4,356 subjects who had data of thyroid function, antithyroid peroxidase antibody (TPOAb), and serum 25-hydroxyvitamin D (25[OH]D) levels. We excluded subjects who were pregnant and who had a history of thyroid disease or thyroid cancer, and those with transient thyroid dysfunction who tested negative for TPOAb (TPOAb[-]). RESULTS: TPOAb positivity (TPOAb[+]) with thyroid dysfunction (subclinical and overt hypothyroidism) was more prevalent in the vitamin D deficient group than in the vitamin D insufficient and sufficient groups including premenopausal (P=0.046) and postmenopausal women (P=0.032), although no significant differences were observed in men. The mean serum 25(OH)D level was significantly lower in the TPOAb(+) with thyroid dysfunction group than in the TPOAb(+) with euthyroidism and TPOAb(-) groups of premenopausal women (P=0.001), although no significant differences were observed in men and postmenopausal women. Multivariate binary logistic regression analysis, adjusted for age, body mass index, and current smoking status, showed that vitamin D insufficiency and deficiency were significantly associated with TPOAb(+) with thyroid dysfunction in premenopausal women (P<0.001), although no significant associations were observed in men and postmenopausal women. CONCLUSION: Low vitamin D status was significantly associated with thyroid autoimmunity and dysfunction in the Korean population, especially in premenopausal women.

Short-term changes in muscle activity and jaw movement patterns after orthognathic surgery in skeletal Class III patients with facial asymmetry.

OBJECTIVE: To evaluate the short-term changes in masticatory muscle activity and mandibular movement patterns after orthognathic surgery in skeletal Class III patients with facial asymmetry. METHODS: Twenty-seven skeletal Class III adult patients were divided into two groups based on the degree of facial asymmetry: the experimental group (n = 17 [11 male and 6 female]; menton deviation ≥ 4 mm) and control group (n = 10 [4 male and 6 female]; menton deviation < 1.6 mm). Cephalography, electromyography (EMG) for the anterior temporalis (TA) and masseter muscles (MM), and mandibular movement (range of motion [ROM] and average chewing pattern [ACP]) were evaluated before (T0) and 7 to 8 months (T1) after the surgery. RESULTS: There were no significant postoperative changes in the EMG potentials of the TA and MM in both groups, except in the anterior cotton roll biting test, in which the masticatory muscle activity had changed into an MM-dominant pattern postoperatively in both groups. In the experimental group, the amount of maximum opening, protrusion, and lateral excursion to the non-deviated side were significantly decreased. The turning point tended to be shorter and significantly moved medially during chewing in the non-deviated side in the experimental group. CONCLUSIONS: In skeletal Class III patients with facial asymmetry, the EMG activity characteristics recovered to presurgical levels within 7 to 8 months after the surgery. Correction of the asymmetry caused limitation in jaw movement in terms of both ROM and ACP on the non-deviated side.

High expression of DEK is associated with poor prognosis in hepatocellular carcinoma.

DEK is an oncogene that has been identified as part of the DEK-CAN fusion gene. DEK plays a role in carcinogenesis through WNT signaling and induces cell proliferation through cyclin-dependent kinase signaling. DEK overexpression has been reported in HCC, but the clinical significance is unclear. This study enrolled 221 cases of HCC. The expression of DEK protein was evaluated by immunohistochemical staining. Cdk4, cyclin D1, Wnt10b, E-cadherin, and ß-catenin were also immunohistochemically stained and analyzed for correlation. The association of clinicopathologic factors with DEK expression was analyzed. DEK expression was observed in 44.8% (99/221) of cases. DEK expression showed a statistical association with clinicopathologic factors, including Edmondson-Steiner grade, presence of vascular emboli, and multiplicity (p<0.05). Among the other IHC markers, the expression of cdk4 was correlated with DEK expression (p<0.05). Patients with high DEK expression showed a significantly lower overall survival rate (p=0.006). However, the disease-free survival rate did not differ significantly. In addition, in a Cox regression model analysis, DEK expression was an independent prognostic factor. In summary, high expression of DEK was observed in HCC and was associated with poor prognostic marker expression and poor prognosis.

Black ginseng activates Akt signaling, thereby enhancing myoblast differentiation and myotube growth.

BACKGROUND: Black ginseng (BG) has greatly enhanced pharmacological activities relative to white or red ginseng. However, the effect and molecular mechanism of BG on muscle growth has not yet been examined. In this study, we investigated whether BG could regulate myoblast differentiation and myotube hypertrophy. METHODS: BG-treated C2C12 myoblasts were differentiated, followed by immunoblotting for myogenic regulators, immunostaining for a muscle marker, myosin heavy chain or immunoprecipitation analysis for myogenic transcription factors. RESULTS: BG treatment of C2C12 cells resulted in the activation of Akt, thereby enhancing heterodimerization of MyoD and E proteins, which in turn promoted muscle-specific gene expression and myoblast differentiation. BG-treated myoblasts formed larger multinucleated myotubes with increased diameter and thickness, accompanied by enhanced Akt/mTOR/p70S6K activation. Furthermore, the BG treatment of human rhabdomyosarcoma cells restored myogenic differentiation. CONCLUSION: BG enhances myoblast differentiation and myotube hypertrophy by activating Akt/mTOR/p70S6k axis. Thus, our study demonstrates that BG has promising potential to treat or prevent muscle loss related to aging or other pathological conditions, such as diabetes.

Optimization of RNA Extraction from Formalin-Fixed Paraffin-Embedded Blocks for Targeted Next-Generation Sequencing.

PURPOSE: Breast cancer has a high prevalence in Korea. To achieve personalized therapy for breast cancer, long-term follow-up specimens are needed for next-generation sequencing (NGS) and multigene analysis. Formalin-fixed paraffin-embedded (FFPE) samples are easier to store than fresh frozen (FF) samples. The objective of this study was to optimize RNA extraction from FFPE blocks for NGS. METHODS: RNA quality from FF and FFPE tissues (n=5), expected RNA amount per unit area, the relationship between archiving time and quantity/quality of FFPE-extracted RNA (n=14), differences in quantitative real-time polymerase chain reaction (qRT-PCR) and NGS results, and comparisons of both techniques with tissue processing at different institutions (n=96) were determined in this study. RESULTS: The quality of RNA did not show any statistically significant difference between paired FF and FFPE specimens (p=0.49). Analysis of tumor cellularity gave an expected RNA amount of 33.25 ng/mm2. Archiving time affected RNA quality, showing a negative correlation with RNA integrity number and a positive correlation with threshold cycle. However, RNA from samples as old as 10 years showed a 100% success rate in qRT-PCR using short primers, showing that the effect of archiving time can be overcome by proper experiment design. NGS showed a higher success rate than qRT-PCR. Specimens from institution B (n=46), which were often stored in a refrigerator for more than 6 hours and fixed without slicing, showed lower success rates and worse results than specimens from the other institutes. CONCLUSION: Archived FFPE tissues can be used to extract RNA for NGS if they are properly processed before fixation. The expected amount of RNA per unit size calculated in this study will be useful for other researchers.

Epithelioid angiomyolipoma of the liver: a case report.

Epithelioid angiomyolipoma (EAML) of liver is a rare neoplasm. Hepatic EAML is often misdiagnosed as other neoplasms such as hepatocellular carcinoma due to non-specific clinical and radiologic features. The morphologic features under microscope and immunohistochemistry staining profile are important in the diagnosis EAML. Here, we report a case of 52-year-old man who found 1.2 cm mass in liver by routine checkup. On the impression of hepatocellular carcinoma, lateral sectionectomy of the liver was done. Microscopically, the tumor is composed of predominant epithelioid cells with vascular component and foamy cells. These cells were positive for HMB45, MelanA, and smooth muscle actin and negative for epithelial membrane antigen. The final diagnosis was hepatic EAML.