Establishment and characterization of six canine hepatocellular carcinoma cell lines.

Background: Hepatocellular carcinoma (HCC) is the most common malignant liver tumor in dogs. Although surgical resection is a major treatment option for canine HCC, there are no distinct strategies for unresectable tumor subtypes or adjuvant chemotherapy for tumors with positive margins. We aimed to establish and characterize novel HCC cell lines from canine patients. Methods: The cellular morphology, general growth features and tumorigenicity of the established cell lines were evaluated. We also examined the sensitivity of the cell lines to multi-target tyrosine kinase inhibitors (TKIs). Results: We established novel canine HCC cell lines from hepatic tumors and an additional kidney tumor of six canine patients. All cell lines showed colony forming and migratory ability. KU-cHCC-001 and KU-cHCC-001-Kidney, two cell lines exhibiting high epithelial-mesenchymal transition characteristics, showed tumorigenicity in xenografted mice. Toceranib, a veterinary TKI that targets vascular endothelial growth factor (VEGFR)/platelet-derived growth factor receptor (PDGFR)/c-kit, effectively inhibited the mitogen-activated protein kinase pathway and induced apoptosis. The established canine HCC cell lines showed greater sensitivity to toceranib than to sorafenib, a first-line treatment for human HCC targeting RAF/VEGFR/PDGFR. Sorafenib showed improved anti-tumor effects when co-treated with SCH772984, an extracellular signal-regulated kinase inhibitor. Conclusion: Our study suggests new therapeutic strategies for canine HCC, and these cell lines are valuable research materials for understanding HCC tumor biology in both humans and dogs.

Sinus irrigation as an adjunctive therapy for odontogenic maxillary sinusitis - an in-depth analysis.

BACKGROUND: Odontogenic maxillary sinusitis (OMS) is widely acknowledged in both the dentistry and otolaryngology fields. Recently, iatrogenic odontogenic maxillary sinusitis cases can be encountered frequently. The purpose of this study was to evaluate the effect of intraoral sinus irrigation using the small lateral window approach in patients with odontogenic maxillary sinusitis by comparing pre- and postoperative volumetric measurement of CBCT and symptoms. We surveyed 21 patients who visited the Oral and Maxillofacial Surgery Department at PNUDH from 2016 to 2022. All the patients' information was extracted from an electronic database. The patients with a follow-up period of 2 months or more were included. The three-dimensional volumetric measurement was performed using the ImageJ program (National Institute of Health, University of Wisconsin). RESULTS: Among 21 patients, 16 (76.1%) were male, and 5 (23%) were female. The most common type of surgery was general anesthesia (16 cases) in which oroantral fistula was present in 7 cases. In the causes of maxillary sinusitis, there were seven implant-related patients, five patients of tooth extraction, seven patients of bone grafting, and two patients in other groups. Radiographic opacity decreased by 40.15% after sinus irrigation especially in bone graft and tooth extraction cases. Clinically, symptoms improved in 17 patients (80.9%). CONCLUSION: By this study, it can be concluded that maxillary sinus irrigation using the small lateral window approach is a clinically and radiologically effective treatment method for odontogenic maxillary sinusitis.

Coagulation Testing in Real-World Setting: Insights From a Comprehensive Survey.

The objective of this survey was to gain a real-world perspective on coagulation testing by evaluating the availability of various coagulation laboratory tests, assessing specific analytic and postanalytic steps in clinical laboratories in Korea.Participants were surveyed using a 65-question questionnaire specifically focused on their coagulation testing practices related to prothrombin time (PT), activated partial thromboplastin time (aPTT), plasma-mixing studies, lupus anticoagulant (LA) tests, platelet function tests, coagulation factor assays, and the composition of hemostasis and thrombosis test panels. The survey was performed between July and September 2022.The survey achieved a 77.9% (81 of 104) response rate. PT or aPTT tests were performed directly at all participating institutions, followed by D-dimer and fibrinogen tests, platelet function test, and plasma-mixing studies in order of frequency. Variations existed in the performance of mixing test and LA assessment. Patterns of coagulating testing differed depending on the size of the hospital. The survey revealed that most laboratories conducted coagulation tests following the international guidelines such as Clinical Laboratory Standards Institute guidelines and the Korean Laboratory Certification system. However, some coagulation tests, including mixing test and LA tests, are yet to be standardized in Korea.Continuous education on coagulation test methods and internal and external quality control are required to encourage laboratories to enhance the performance of coagulation testing.

Clinical application of prospective whole-exome sequencing in the diagnosis of genetic disease: Experience of a regional disease center in South Korea.

INTRODUCTION: Next-generation sequencing helps clinicians diagnose patients with suspected genetic disorders. The current study aimed to investigate the diagnostic yield and clinical utility of prospective whole-exome sequencing (WES) in rare diseases. METHODS: WES was performed in 92 patients who presented with clinical symptoms suggestive of genetic disorders. The WES data were analyzed using an in-house developed software. The patients' phenotypic characteristics were classified according to the human phenotype ontology. RESULTS: WES detected 64 variants, 13 were classified as pathogenic, 26 as likely pathogenic, and 25 as variants of uncertain significance. In 57 patients with these variants, 30 were identified as causal variants. The diagnostic yield was higher in patients with abnormalities in joint mobility and skin morphology than in those with cerebellar hypoplasia/atrophy, epilepsy, global developmental delay, dysmorphic features/facial dysmorphisms, and chronic kidney disease/abnormal renal morphology. CONCLUSION: In this study, a WES-based variant interpretation system was employed to provide a definitive diagnosis for 28.3% of the patients suspected of having genetic disorders. WES is particularly useful for diagnosing rare diseases with symptoms that affect more than one system, when targeted genetic panels are difficult to employ.

Delirium After Traumatic Brain Injury: Prediction by Location and Size of Brain Lesion.

OBJECTIVE: To examine (1) the location of brain lesion that would predict post-traumatic delirium and (2) the association between volume of brain lesion and occurrence of delirium in patients with traumatic brain injury (TBI). METHODS: A retrospective study was conducted by reviewing medical records of 68 TBI patients, categorized into two groups: the delirious group (n=38) and non-delirious group (n=30). The location and volume of TBI were investigated with the 3D Slicer software. RESULTS: The TBI region in the delirious group mainly involved the frontal or temporal lobe (p=0.038). All 36 delirious patients had brain injury on the right side (p=0.046). The volume of hemorrhage in the delirious group was larger by about 95 mL compared to the non-delirious group, but this difference was not statistically significant (p=0.382). CONCLUSION: Patients with delirium after TBI had significantly different injury site and side, but not lesion size compared to patients without delirium.

Oncogenic fusion of BCAR4 activates EGFR signaling and is sensitive to dual inhibition of EGFR/HER2.

We previously reported CD63-BCAR4 fusion as a novel oncogene that significantly enhanced cell migration and metastasis in lung cancer. To identify effective inhibitors of metastatic activity induced by BCAR4 fusion, we screened a drug library of 381 FDA-approved compounds. The effect of drugs on cell migration was evaluated by monitoring wound healing. Drugs that decreased the cellular mobility of fusion-overexpressing cells compared with that of control cells were selected as candidates. Library screening revealed that erlotinib, canertinib, and lapatinib demonstrated inhibitory effects on cell migration. Activation of the EGFR signaling pathway was detected after ectopic expression of CD63-BCAR4 in normal bronchial epithelial cells, as observed by the increased phosphorylation of tyrosine residues in the EGFR protein. We also confirmed increased levels of the phosphorylated EGFR protein in resected tumors from mice injected with CD63-BCAR4 overexpressing cells. Tyrosine kinase inhibitors (TKIs) of the EGFR family significantly inhibit the migration of BCAR4 fusion-overexpressing cells and induce apoptosis at high concentrations. Among the EGFR family TKIs, canertinib, a dual EGFR/HER2 inhibitor, showed the best inhibitory effect on the migration and viability of BCAR4 fusion-overexpressing cells. We examined the effect of canertinib in vivo using a mouse xenograft model. Oral administration of canertinib to xenografted mice reduced tumor growth induced by the CD63-BCAR4 fusion gene. In addition, canertinib treatment restored E-cadherin expression and reduced the expression of epithelial-mesenchymal transition regulatory factors such as Slug and Snail. Taken together, these results suggest that EGFR/HER2 inhibitors are potential therapeutic options for BCAR4 fusion-harboring lung cancer patients, even in the absence of EGFR mutations.

Establishment of Canine Transitional Cell Carcinoma Cell Lines Harboring BRAF V595E Mutation as a Therapeutic Target.

Transitional cell carcinoma (TCC) is the most common malignant tumor of the canine urinary tract and tends to have a poor prognosis due to its invasive potential. Recent studies have reported that up to 80% of canine urothelial carcinoma has the BRAF V595E mutation, which is homologous to the human V600E mutation. Activating the BRAF mutation is an actionable target for developing effective therapeutic agents inhibiting the BRAF/mitogen-activated protein kinase (MAPK) pathway in canine cancer as well as human cancer. We established novel canine TCC cell lines from two tumor tissues and one metastatic lymph node of canine TCC patients harboring the BRAF V595E mutation. Tumor tissues highly expressed the BRAF mutant and phosphorylated extracellular signal-related kinases (ERK)1/2 proteins. The derived cell lines demonstrated activated MAPK pathways. We also evaluated the cell lines for sensitivity to BRAF inhibitors. Sorafenib, a multiple kinase inhibitor targeting RAF/vascular endothelial growth factor receptor (VEGFR), successfully inhibited the BRAF/MAPK pathway and induced apoptosis. The established canine TCC cell lines responded with greater sensitivity to sorafenib than to vemurafenib, which is known as a specific BRAF inhibitor in human cancer. Our results demonstrated that canine TCC cells showed different responses compared to human cancer with the BRAF V600E mutation. These cell lines would be valuable research materials to develop therapeutic strategies for canine TCC patients.

12α-Hydroxylated bile acid induces hepatic steatosis with dysbiosis in rats.

There is an increasing need to explore the mechanism of the progression of non-alcoholic fatty liver disease. Steroid metabolism is closely linked to hepatic steatosis and steroids are excreted as bile acids (BAs). Here, we demonstrated that feeding WKAH/HkmSlc inbred rats a diet supplemented with cholic acid (CA) at 0.5 g/kg for 13 weeks induced simple steatosis without obesity. Liver triglyceride and cholesterol levels were increased accompanied by mild elevation of aminotransferase activities. There were no signs of inflammation, insulin resistance, oxidative stress, or fibrosis. CA supplementation increased levels of CA and taurocholic acid (TCA) in enterohepatic circulation and deoxycholic acid (DCA) levels in cecum with an increased ratio of 12α-hydroxylated BAs to non-12α-hydroxylated BAs. Analyses of hepatic gene expression revealed no apparent feedback control of BA and cholesterol biosynthesis. CA feeding induced dysbiosis in cecal microbiota with enrichment of DCA producers, which underlines the increased cecal DCA levels. The mechanism of steatosis was increased expression of Srebp1 (positive regulator of liver lipogenesis) through activation of the liver X receptor by increased oxysterols in the CA-fed rats, especially 4ß-hydroxycholesterol (4ßOH) formed by upregulated expression of hepatic Cyp3a2, responsible for 4ßOH formation. Multiple regression analyses identified portal TCA and cecal DCA as positive predictors for liver 4ßOH levels. The possible mechanisms linking these predictors and upregulated expression of Cyp3a2 are discussed. Overall, our observations highlight the role of 12α-hydroxylated BAs in triggering liver lipogenesis and allow us to explore the mechanisms of hepatic steatosis onset, focusing on cholesterol and BA metabolism.

Performance evaluation of the ZL 6000i cone-plate rotational viscometer.

BACKGROUND: Elevated whole blood viscosity (WBV) is associated with increasing blood flow resistance in the microcirculation and is used to predict the occurrence and outcome of cardiovascular diseases. OBJECTIVE: We evaluated the analytical performance of the ZL 6000i (Zonci Technology, Beijing, China), new fully automated cone-plate rotational type viscometer. METHODS: We collected blood samples from 287 adults to establish a reference range. We evaluated total precision for 20 days using quality control (QC) samples and within-run precision was evaluated by performing 50 consecutive measurements of blood samples. Aliquots of blood samples were stored at room temperature and 5°C to assess the effect of storage time and temperature on viscosity measurement. RESULTS: The reference ranges and median values of WBV were significantly higher in males than in females due to difference in hematocrit level. The coefficients of variation (CVs) for precision using QC and human blood samples were < 7.5% at all shear rates. WBV of the samples stored at room temperature for up to 6 h, and at 5°C for samples stored up to 2 days was not different from the control. CONCLUSIONS: In this study, the ZL 6000i viscometer yields reproducible WBV data and is clinically useful for monitoring viscosity.

15-Keto prostaglandin E2 induces heme oxygenase-1 expression through activation of Nrf2 in human colon epithelial CCD 841 CoN cells.

Prostaglandin E2 (PGE2) plays a key role in inflammation-associated carcinogenesis. NAD+-dependent 15-hydroxyprostaglandin dehydrogenase (15-PGDH) catalyzes the oxidation of the 15(S)-hydroxyl group of PGE2 to generate 15-keto PGE2. 15-PGDH has been known as a tumor suppressor in various malignancies including colon cancer. However, the molecular mechanisms underlying the tumor-suppressive function of 15-PGDH remain largely unresolved. In this study, we found that 15-keto PGE2 upregulated the expression of heme oxygenase-1 (HO-1), a representative antioxidative and anti-inflammatory enzyme, at both transcriptional and translational levels, in human colon epithelial CCD 841 CoN cells. A redox-sensitive transcription factor, NF-E2-related factor (Nrf2) plays a critical role in the regulation of HO-1 and other cytoprotective proteins. 15-Keto PGE2 induced translocation of Nrf2 into the nucleus and antioxidant response element-driven luciferase activity. Furthermore, the silencing of the Nrf2 gene abolished 15-keto PGE2-induced HO-1 expression in CCD 841 CoN cells. 15-Keto PGE2 activated AKT signaling, and the pharmacological AKT inhibitor, LY294002 suppressed the 15-keto PGE2-induced HO-1 expression. 15-Keto PGE2 generates the reactive oxygen species which is suppressed by the general antioxidant N-acetyl-l-cysteine. N-acetyl-l-cysteine treatment attenuated the 15-keto PGE2-induced phosphorylation of GSK3ß, transcriptional activity of Nrf2, and subsequently HO-1 expression. However, 13,14-dihydro-15-keto PGE2 lacking the α,ß-unsaturated carbonyl moiety failed to induce intracellular production of reactive oxygen species, HO-1 expression and nuclear translocation of Nrf2. In conclusion, 15-keto PGE2 induces HO-1 expression through Nrf2 activation in human colon epithelial cells.

Relationship between Alcohol Consumption and Ocular Pressure according to Facial Flushing in Korean Men with Obesity.

BACKGROUND: This study aimed to examine the relationship between alcohol consumption and intraocular pressure (IOP) according to facial flushing in Korean men with obesity. METHODS: The study included 479 Korean men with a body mass index of ≥25 kg/m2 (75 non-drinkers, 174 with drinking-related facial flushing, and 230 without facial flushing) who underwent health check-ups between October 1, 2016 and March 31, 2017. Multivariate logistic regression was used to assess the relationship between alcohol consumption and high IOP (≥21 mm Hg). RESULTS: Flushers consuming ≤16 drinks per week had a significantly higher risk of high IOP than non-drinkers, depending on alcohol consumption (≤8 standard drinks: odds ratio [OR], 4.49; 95% confidence interval [CI], 1.05- 19.25; >8 but ≤16 standard drinks: OR, 8.14; 95% CI, 1.37-48.45). However, when the consumption was >16 drinks per week, the high IOP risk did not significantly increase (OR, 0.71; 95% CI, 0.05-10.69). In addition, there was no significant relationship between alcohol consumption and high IOP among non-flushers consuming ≤8 drinks per week (OR, 2.07; 95% CI, 0.52-8.19). However, a significantly increased risk of high IOP was observed among nonflushers consuming >8 drinks per week, depending on alcohol consumption (>8 but ≤16 standard drinks: OR, 4.84; 95% CI, 1.14-20.61; >16 standard drinks: OR, 4.08; 95% CI, 1.02-16.26). CONCLUSION: This study suggests that obese men with alcohol flush reactions may have an increased risk of high IOP with the consumption of smaller amounts of alcohol than non-flushers.

Inhibitory Effect of Olive Leaf Extract on Obesity in High-fat Diet-induced Mice.

BACKGROUND/AIM: The rapid increase in the number of people who are overweight or obese, which increases the risk of diseases and health problems, is becoming an important issue. Herein, we investigated whether olive leaf extract (OLE) has potent anti-obesity effects in high-fat induced mouse models. MATERIALS AND METHODS: C57BL/6 mice were randomized into normal control, high-fat diet (HFD), HFD with OLE, and HFD with garcinia groups and administered experimental diets for 12 weeks. Body weight and food intake were measured once per week and obesity-related biomarkers were evaluated in the serum and adipose tissue. RESULTS: OLE significantly suppressed weight gain, food efficiency ratio, visceral fat accumulation, and serum lipid composition in HFD-induced mice. Furthermore, the expression of adipogenesis- and thermogenesis-related molecules was decreased in the OLE-treated group. CONCLUSION: OLE prevents obesity development by regulating the expression of molecules involved in adipogenesis and thermogenesis.

15-Deoxy-Δ12,14-prostaglandin J2 up-regulates the expression of 15-hydroxyprostaglandin dehydrogenase through DNA methyltransferase 1 inactivation.

15-Hydroxyprostaglandin dehydrogenase (15-PGDH) is the key enzyme that catalyses the conversion of prostaglandin E2 to a keto metabolite. The expression of 15-PGDH is ubiquitously repressed in various human malignancies. However, the molecular mechanisms underlying down-regulation of 15-PGDH expression remain largely unknown. 15-Deoxy-△12,14-prostaglandin J2 (15d-PGJ2), an endogenous ligand of peroxisome proliferator-activated receptor γ, has been reported to have anti-inflammatory and anticarcinogenic activities. In the present study, we have found that 15d-PGJ2 induces expression and catalytic activity of 15-PGDH in human breast cancer (MDA-MB-231) cells. 15d-PGJ2 decreased the level of CpG methylation in the 15-PGDH promoter in MDA-MB-231 cells as determined by the bisulphite genome sequencing and methyl-specific PCR. 15d-PGJ2 inhibited the catalytic activity of methyltransferase 1 (DNMT1) but did not influence its expression. Biotinylated 15d-PGJ2 directly interacted with DNMT1 and reduced its catalytic activity. Chromatin-immunoprecipitation analysis revealed that 15d-PGJ2 significantly attenuated DNMT1 binding to the activator protein-1 transcription factor present in the 15-PGDH promoter region. A nonelectrophilic analogue 9,10-dihydro-15d-PGJ2 failed to suppress the methylation of CpG islands present in 15-PGDH promoter and did not affect both DNMT1 activity and 15-PGDH expression. These findings suggest that the α,ß-unsaturated carbonyl group present in 15d-PGJ2 is essential for its inactivation on DNMT1 and expression of 15-PGDH. In conclusion, 15d-PGJ2 plays as a hypomethylating agent through direct interaction with DNMT1 and consequently suppresses DNMT1-mediated hypermethylation of 15-PGDH promoter, leading to up-regulation of 15-PGDH expression.

A mitochondrial DNA D loop insertion detected almost exclusively in non-replicating tissues with maternal inheritance across three generations.

Muscle biopsy identified a possibly pathogenic, mitochondrial DNA D-loop insertion, in each of 5 family members from two generations, that was otherwise undetectable in most other tissues. The tissue specific regulation of heteroplasmy is reflected in an age related increase in muscle heteroplasmy level, across the pedigree. This latter finding is in keeping with previous reports (e.g. T408A, C16327) but differs in having a very high muscle heteroplasmy level, and appears maternally transmitted.

How safe is tuning a radio?: using the radio tuning task as a benchmark for distracted driving.

Drivers engage in non-driving tasks while driving, such as interactions entertainment systems. Studies have identified glance patterns related to such interactions, and manual radio tuning has been used as a reference task to set an upper bound on the acceptable demand of interactions. Consequently, some view the risk associated with radio tuning as defining the upper limit of glance measures associated with visual-manual in-vehicle activities. However, we have little knowledge about the actual degree of crash risk that radio tuning poses and, by extension, the risk of tasks that have similar glance patterns as the radio tuning task. In the current study, we use counterfactual simulation to take the glance patterns for manual radio tuning tasks from an on-road experiment and apply these patterns to lead-vehicle events observed in naturalistic driving studies. We then quantify how often the glance patterns from radio tuning are associated with rear-end crashes, compared to driving only situations. We used the pre-crash kinematics from 34 crash events from the SHRP2 naturalistic driving study to investigate the effect of radio tuning in crash-imminent situations, and we also investigated the effect of radio tuning on 2,475 routine braking events from the Safety Pilot project. The counterfactual simulation showed that off-road glances transform some near-crashes that could have been avoided into crashes, and glance patterns observed in on-road radio tuning experiment produced 2.85-5.00 times more crashes than baseline driving.

Tumour size, volume, and marker expression during radiation therapy can predict survival of cervical cancer patients: a multi-institutional retrospective analysis of KROG 16-01.

OBJECTIVE: The aim of this multi-institutional study was to determine the prognostic impact of tumour parameters, such as tumour size (TS), tumour volume (TV), and marker expression, on survival during radiation therapy (RT) for cervical cancer patients. METHODS: A total of 231 patients with histologically confirmed cervical cancer, classified as Federation of Gynecology and Obstetrics (FIGO) Ib2-IVa, were enrolled in this study. Pre- and mid-RT pelvic magnetic resonance imaging (MRI) and squamous cell carcinoma antigen (SCC-ag) analysis were performed twice, during RT and just before brachytherapy. RESULTS: The median follow-up time was 27.8months (range, 2-116months). Multivariate analysis revealed that stage (odds ratio [OR], 2.936 and 95% confidence interval [CI], 1.119-7.707; P=0.029), tumour volume reduction rate (TVRR) (OR, 3.435 and 95% CI, 1.062-11.106; P=0.039), and SCC-ag reduction rate (SCCRR) (OR, 5.104 and 95% CI, 1.769-14.727; P=0.003) were independently associated with overall survival (OS), while pre-RT TS (OR, 2.148 and 95% CI, 1.221-3.810; P=0.009), mid-RT TV (OR, 3.106 and 95% CI, 1.685-5.724; P<0.0001) and SCCRR (OR, 1.954 and 95% CI, 1.133-3.369; P=0.016) were associated with progression-free survival (PFS). Based on the prognostic factor analysis, patients with the highest prognostic risk score of 3 showed poorer overall survival and progression free survival than patients with lower prognostic risk scores. CONCLUSION: We identified that tumour parameters such as TVRR, SCCRR, pre-RT TS, and mid-RT TV areindependent and strong prognostic parameters for patients with cervical cancer receiving RT. This scoring system-based prognostic factor analysis could be used to help develop optimized treatment plans for cervical cancer patients during RT.

Chemopreventive and Chemotherapeutic Effects of Fish Oil derived Omega-3 Polyunsaturated Fatty Acids on Colon Carcinogenesis.

Colorectal cancer is the third most common cause of cancer related death in the world. Multiple lines of evidence suggest that there is an association between consumption of dietary fat and colon cancer risk. Not only the amount but also the type and the ratio of fatty acids comprising dietary fats consumed have been implicated in the etiology and pathogenesis of colon cancer. Omega-3 (n-3) polyunsaturated fatty acids (PUFAs), such as docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), have been known to inhibit development of colon cancer by downregulating the expression of genes involved in colon carcinogenesis and also by altering the membrane lipid composition. Data from laboratory, epidemiological, and clinical studies substantiate the beneficial role of n-3 PUFAs in preventing colitis and subsequent development of colon cancer. In addition, recent studies suggest that some n-3 PUFAs can be effective as an adjuvant with chemotherapeutic agents and other natural anticancer compounds in the management of colon cancer. In this review, we discuss chemopreventive and therapeutic effects of fish oil derived long chain n-3 PUFAs, particularly EPA and DHA, with focus on synergetic effects of which they exert when combined with chemotherapeutic agents and other natural compounds.