LCK-Mediated RIPK3 Activation Controls Double-Positive Thymocyte Proliferation and Restrains Thymic Lymphoma by Regulating the PP2A-ERK Axis.

Receptor-interacting protein kinase 3 (RIPK3) is the primary regulator of necroptotic cell death. RIPK3 expression is often silenced in various cancer cells, which suggests that it may have tumor suppressor properties. However, the exact mechanism by which RIPK3 negatively regulates cancer development and progression remains unclear. This report indicates that RIPK3 acts as a potent regulator of the homeostatic proliferation of CD4+ CD8+ double-positive (DP) thymocytes. Abnormal proliferation of RIPK3-deficient DP thymocytes occurs independently of the well-known role for RIPK3 in necroptosis (upstream of MLKL activation), and is associated with an incidental thymic mass, likely thymic hyperplasia. In addition, Ripk3-null mice develop increased thymic tumor formation accompanied by reduced host survival in the context of an N-ethyl-N-nitrosourea (ENU)-induced tumor model. Moreover, RIPK3 deficiency in p53-null mice promotes thymic lymphoma development via upregulated extracellular signal-regulated kinase (ERK) signaling, which correlates with markedly reduced survival rates. Mechanistically, lymphocyte-specific protein tyrosine kinase (LCK) activates RIPK3, which in turn leads to increases in the phosphatase activity of protein phosphatase 2 (PP2A), thereby suppressing hyper-activation of ERK in DP thymocytes. Overall, these findings suggest that a RIPK3-PP2A-ERK signaling axis regulates DP thymocyte homeostasis and may provide a potential therapeutic target to improve thymic lymphoma therapies.

Sex-Biased Molecular Signature for Overall Survival of Liver Cancer Patients.

Sex/gender disparity has been shown in the incidence and prognosis of many types of diseases, probably due to differences in genes, physiological conditions such as hormones, and lifestyle between the sexes. The mortality and survival rates of many cancers, especially liver cancer, differ between men and women. Due to the pronounced sex/gender disparity, considering sex/ gender may be necessary for the diagnosis and treatment of liver cancer. By analyzing research articles through a PubMed literature search, the present review identified 12 genes which showed practical relevance to cancer and sex disparities. Among the 12 sex-specific genes, 7 genes (BAP1, CTNNB1, FOXA1, GSTO1, GSTP1, IL6, and SRPK1) showed sex-biased function in liver cancer. Here we summarized previous findings of cancer molecular signature including our own analysis, and showed that sexbiased molecular signature CTNNB1High, IL6High, RHOAHigh and GLIPR1Low may serve as a female-specific index for prediction and evaluation of OS in liver cancer patients. This review suggests a potential implication of sex-biased molecular signature in liver cancer, providing a useful information on diagnosis and prediction of disease progression based on gender.

TRIM24-RIP3 axis perturbation accelerates osteoarthritis pathogenesis.

OBJECTIVES: Recently, necroptosis has attracted increasing attention in arthritis research; however, it remains unclear whether its regulation is involved in osteoarthritis (OA) pathogenesis. Since receptor-interacting protein kinase-3 (RIP3) plays a pivotal role in necroptosis and its dysregulation is involved in various pathological processes, we investigated the role of the RIP3 axis in OA pathogenesis. METHODS: Experimental OA was induced in wild-type or Rip3 knockout mice by surgery to destabilise the medial meniscus (DMM) or the intra-articular injection of adenovirus carrying a target gene (Ad-Rip3 and Ad-Trim24 shRNA). RIP3 expression was examined in OA cartilage from human patients; Trim24, a negative regulator of RIP3, was identified by microarray and in silico analysis. Connectivity map (CMap) and in silico binding approaches were used to identify RIP3 inhibitors and to examine their direct regulation of RIP3 activation in OA pathogenesis. RESULTS: RIP3 expression was markedly higher in damaged cartilage from patients with OA than in undamaged cartilage. In the mouse model, adenoviral RIP3 overexpression accelerated cartilage disruption, whereas Rip3 depletion reduced DMM-induced OA pathogenesis. Additionally, TRIM24 knockdown upregulated RIP3 expression; its downregulation promoted OA pathogenesis in knee joint tissues. The CMap approach and in silico binding assay identified AZ-628 as a potent RIP3 inhibitor and demonstrated that it abolished RIP3-mediated OA pathogenesis by inhibiting RIP3 kinase activity. CONCLUSIONS: TRIM24-RIP3 axis perturbation promotes OA chronicity by activating RIP3 kinase, suggesting that the therapeutic manipulation of this pathway could provide new avenues for treating OA.

Spheroid-Induced Epithelial-Mesenchymal Transition Provokes Global Alterations of Breast Cancer Lipidome: A Multi-Layered Omics Analysis.

Metabolic rewiring has been recognized as an important feature to the progression of cancer. However, the essential components and functions of lipid metabolic networks in breast cancer progression are not fully understood. In this study, we investigated the roles of altered lipid metabolism in the malignant phenotype of breast cancer. Using a spheroid-induced epithelial-mesenchymal transition (EMT) model, we conducted multi-layered lipidomic and transcriptomic analysis to comprehensively describe the rewiring of the breast cancer lipidome during the malignant transformation. A tremendous homeostatic disturbance of various complex lipid species including ceramide, sphingomyelin, ether-linked phosphatidylcholines, and ether-linked phosphatidylethanolamine was found in the mesenchymal state of cancer cells. Noticeably, polyunsaturated fatty acids composition in spheroid cells was significantly decreased, accordingly with the gene expression patterns observed in the transcriptomic analysis of associated regulators. For instance, the up-regulation of SCD, ACOX3, and FADS1 and the down-regulation of PTPLB, PECR, and ELOVL2 were found among other lipid metabolic regulators. Significantly, the ratio of C22:6n3 (docosahexaenoic acid, DHA) to C22:5n3 was dramatically reduced in spheroid cells analogously to the down-regulation of ELOVL2. Following mechanistic study confirmed the up-regulation of SCD and down-regulation of PTPLB, PECR, ELOVL2, and ELOVL3 in the spheroid cells. Furthermore, the depletion of ELOVL2 induced metastatic characteristics in breast cancer cells via the SREBPs axis. A subsequent large-scale analysis using 51 breast cancer cell lines demonstrated the reduced expression of ELOVL2 in basal-like phenotypes. Breast cancer patients with low ELOVL2 expression exhibited poor prognoses (HR = 0.76, CI = 0.67-0.86). Collectively, ELOVL2 expression is associated with the malignant phenotypes and appear to be a novel prognostic biomarker in breast cancer. In conclusion, the present study demonstrates that there is a global alteration of the lipid composition during EMT and suggests the down-regulation of ELOVL2 induces lipid metabolism reprogramming in breast cancer and contributes to their malignant phenotypes.

Comparison of Clinical Findings of Congenital Muscular Torticollis Between Patients With and Without Sternocleidomastoid Lesions as Determined by Ultrasonography.

AIM: To compare clinical findings for patients with congenital muscular torticollis (CMT) between those with and without a sternocleidomastoid (SCM) lesion. METHODS: Medical records of 182 patients with CMT were retrospectively reviewed and the patients were divided into SCM lesion and nonlesion groups by ultrasonographic results. Intrauterine position, age, duration of therapy, rotation/tilting side, and the passive range of motion and angle of the neck were compared. RESULTS: There were 74 SCM lesion and 108 nonlesion cases. The mean age at the first visit was 55.3 days in the SCM lesion group and 146.6 days in the nonlesion group. The mean therapy time in the nonlesion group was 66.5 days, significantly shorter than for the SCM lesion group (117.5 d). Tilting and rotation of the head in the same direction was observed only in the nonlesion group (n=9, 8.3%). Rotational limitation of the affected muscle side was 22.6 degree in the SCM lesion and 3.6 degree in the nonlesion group, and the tilting limitation was 19.2 degree in the SCM lesion and 10.4 degree in the nonlesion group. CONCLUSIONS: The nonlesion group had a better prognosis with shorter treatment duration. This group was more limited in head tilting than in head rotation, and the pattern of head rotation/tilting in the same direction was observed only in this group. These findings suggest that pathophysiological mechanisms and clinical characteristics may differ between CMT patients with and without SCM lesions. LEVEL OF EVIDENCE: Level II-prognostic studies, retrospective study.

A formulated red ginseng extract inhibits autophagic flux and sensitizes to doxorubicin-induced cell death.

BACKGROUND: Ginseng is believed to have antitumor activity. Autophagy is largely a prosurvival cellular process that is activated in response to cellular stressors, including cytotoxic chemotherapy; therefore, agents that inhibit autophagy can be used as chemosensitizers in cancer treatment. We examined the ability of Korean Red Ginseng extract (RGE) to prevent autophagic flux and to make hepatocellular carcinoma (HCC) cells become more sensitive to doxorubicin. METHODS: The cytotoxic effects of total RGE or its saponin fraction (RGS) on HCC cells were examined by the lactate dehydrogenase assay in a dose- or time-dependent manner. The effect of RGE or RGS on autophagy was measured by analyzing microtubule-associated protein 1A/1B-light chain (LC)3-II expression and LC3 puncta formation in HCC cells. Late-stage autophagy suppression was tested using tandem-labeled green fluorescent protein (GFP)-monomeric red fluorescent protein (mRFP)-LC3. RESULTS: RGE markedly increased the amount of LC3-II, but green and red puncta in tandem-labeled GFP-mRFP-LC3 remained colocalized over time, indicating that RGE inhibited autophagy at a late stage. Suppression of autophagy through knockdown of key ATG genes increased doxorubicin-induced cell death, suggesting that autophagy induced by doxorubicin has a protective function in HCC. Finally, RGE and RGS markedly sensitized HCC cells, (but not normal liver cells), to doxorubicin-induced cell death. CONCLUSION: Our data suggest that inhibition of late-stage autophagic flux by RGE is important for its potentiation of doxorubicin-induced cancer cell death. Therapy combining RGE with doxorubicin could serve as an effective strategy in the treatment of HCC.

PELI1 Selectively Targets Kinase-Active RIP3 for Ubiquitylation-Dependent Proteasomal Degradation.

Receptor-interacting protein kinase-3 (RIP3 or RIPK3) is a central protein in necroptosis, but posttranslational processes that regulate RIP3 activity and stability remain poorly understood. Here, we identify pellino E3 ubiquitin protein ligase 1 (PELI1) as an E3 ligase that targets RIP3 for proteasome-dependent degradation. Phosphorylation of RIP3 on T182 leads to interaction with the forkhead-associated (FHA) domain of PELI1 and PELI1-mediated K48-linked polyubiquitylation of RIP3 on K363. This same phosphorylation event is also important for RIP3 kinase activity; thus, PELI1 preferentially targets kinase-active RIP3 for degradation. PELI1-mediated RIP3 degradation effectively prevents cell death triggered by RIP3 hyperactivation. Importantly, upregulated RIP3 expression in keratinocytes from toxic epidermal necrolysis (TEN) patients is correlated with low expression of PELI1, suggesting that loss of PELI1 may play a role in the pathogenesis of TEN. We propose that PELI1 may function to control inadvertent activation of RIP3, thus preventing aberrant cell death and maintaining cellular homeostasis.

TLR4 Endogenous Ligand S100A8/A9 Levels in Adult-Onset Still's Disease and Their Association with Disease Activity and Clinical Manifestations.

S100A8/A9 has been suggested as a marker of disease activity in patients with adult-onset Still's disease (AOSD). We evaluated the clinical significance of S100A8/A9 as a biomarker and its pathogenic role in AOSD. Blood samples were collected prospectively from 20 AOSD patients and 20 healthy controls (HCs). Furthermore, skin and lymph node biopsy specimens of AOSD patients were investigated for S100A8/A9 expression levels via immunohistochemistry. Peripheral blood mononuclear cells (PBMCs) of active AOSD patients and HCs were investigated for S100A8/A9 cell signals. S100A8/A9, interleukin-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α) levels in active AOSD patients were higher than those of HCs. S100A8/A9 levels correlated positively with IL-1ß, TNF-α and C-reactive protein. The inflammatory cells expressing S100A8/A9 were graded from one to three in skin and lymph node biopsies of AOSD patients. The grading for S100A8/A9 was more intense in the skin lesions with karyorrhexis, mucin deposition, and neutrophil infiltration. Like lipopolysaccharide (LPS), S100A8/A9 induced phosphorylation of p38 and c-Jun amino-terminal kinase (JNK) in PBMCs, suggesting that S100A8/A9 activates Toll-like receptor 4 signaling pathways. These findings suggest that S100A8/A9 may be involved in the inflammatory response with induction of proinflammatory cytokines and may serve as a clinicopathological marker for disease activity in AOSD.

Prediction of Early Recurrence After Acute Ischemic Stroke.

IMPORTANCE: Approximately half of recurrent strokes occur within days and weeks of an ischemic stroke. It is imperative to identify patients at imminent risk of recurrent stroke because recurrent events lead to prolonged hospitalization, worsened functional outcome, and increased mortality. OBJECTIVE: To test the validity of a prognostic score that was exclusively developed to predict early risk of stroke recurrence in a multicenter setting. DESIGN, SETTING, AND PARTICIPANTS: This hospital-based cohort study examined patients with and without magnetic resonance imaging-confirmed recurrent stroke within 90 days after an ischemic stroke. The study was performed at 3 teaching hospitals in the United States, Brazil, and South Korea and comprised adult patients admitted within 72 hours of symptom onset with a magnetic resonance imaging-confirmed diagnosis of acute ischemic stroke. Recruitment to the US cohort was performed from June 1, 2009, through April 30, 2011. Recruitment to the Korean and Brazilian cohorts was performed from January 1, 2007, through December 31, 2011. Data analysis was performed from June 1, 2013, to December 31, 2014. MAIN OUTCOMES AND MEASURES: The primary outcome was recurrent ischemic stroke as defined by a clinical incident that was clearly attributable to a new area of brain infarction occurring within the 90 days of index infarction. An investigator who was masked to the patient's recurrence status calculated the Recurrence Risk Estimator (RRE) score for each patient based on information available after initial line of testing in the emergency department. We assessed the predictive performance of the RRE by computing the area under the receiver operating characteristic curve. RESULTS: The study included 1468 consecutive patients with 59 recurrent ischemic stroke events. The median age of the patients was 69 (interquartile range, 58-79) years, and 633 (43.1%) were female. The cumulative 90-day recurrence rate was 4.2% (95% CI, 3.2%-5.2%). The mean RRE score was 2.2 (95% CI, 1.9-2.5) in patients with recurrence and 1.0 (95% CI, 1.0-1.1) in patients without. The risk of recurrence increased with a higher RRE score (log-rank test, P < .001). The area under the receiver operating characteristic curve for discrimination was 0.76 (95% CI, 0.70-0.82). The RRE identified 710 patients (48.4%) in the study population as high risk (>10%) or low risk (<1%). The sensitivity and specificity were 38% and 93% for identifying low-risk subsets and 41% and 90% for identifying high-risk subsets, respectively. CONCLUSIONS AND RELEVANCE: This study confirms the validity of the RRE score in a multicenter cohort of patients with diverse characteristics. Our findings suggest that the RRE could be useful in identifying high- and low-risk patients for targeted stroke prevention.

Anti-septic effects of dabrafenib on HMGB1-mediated inflammatory responses.

A nucleosomal protein, high mobility group box 1 (HMGB1) is known to be a late mediator of sepsis. Dabrafenib is a B-Raf inhibitor and initially used for the treatment of metastatic melanoma therapy. Inhibition of HMGB1 and renewal of vascular integrity is appearing as an engaging therapeutic strategy in the administration of severe sepsis or septic shock. Here, we examined the effects of dabrafenib (DAB) on the modulation of HMGB1-mediated septic responses. DAB inhibited the release of HMGB1 and downregulated HMGB1-dependent inflammatory responses by enhancing the expressions of cell adhesion molecules (CAMs) in human endothelial cells. In addition, treatment with DAB inhibited the HMGB1 secretion by CLP and sepsis-related mortality and pulmonary injury. This study demonstrated that DAB could be alternative therapeutic options for sepsis or septic shock via the inhibition of the HMGB1 signaling pathway. [BMB Reports 2016; 49(4): 214-219].

Long term outcome and predictors of ischemic stroke recurrence in adult moyamoya disease.

OBJECTIVE: We sought to identify the clinical outcome and predictors for ischemic stroke recurrence in adults with symptomatic moyamoya disease (MMD). METHODS: We analyzed 104 adult MMD patients with ischemic stroke or TIA registered at our institution. All patients underwent digital subtraction angiography and single photon emission computed tomography to measure disease severity and cerebral vascular reserve (CVR). A Cox regression model was used to identify predictors of recurrent ischemic stroke. RESULTS: Fifty-nine patients were non-surgically treated and 45 patients were surgically treated. In the non-surgical group, the Kaplan-Meier estimate of ischemic stroke recurrence was 1.6% in the first year and 11.8% in the 5th year. Hypertension (hazard ratio [HR]=0.07, 95% confidence interval [CI] 0.01-0.99), diabetes (HR=35.16, 95% CI 2.61-474.16), presence of steno-occlusive lesion in posterior cerebral arteries (HR=17.53, 95% CI 2.02-152.43), and extended or global decreased CVR (HR=13.62, 95% CI 1.55-119.84) were independent predictors of recurrence. In the surgical group, the Kaplan-Meier estimate of ischemic stroke recurrence was 24.4% in the first year and 24.4% in the 5th year. Half of the recurred patients experienced recurrent ischemic strokes postoperatively. Diabetes was the only predictor of recurrent ischemic stroke (HR=6.17, 95% CI 1.31-29.14). CONCLUSIONS: In non-surgically treated MMD, PCA stenosis and CVR were identified as predictors of ischemic stroke recurrence. Diabetes was an independent predictor of recurrent ischemic stroke in both non-surgical and surgically treated MMD groups.

Gastrointestinal stromal tumor solitary distant recurrence in the left brachialis muscle.

Gastrointestinal stromal tumors (GISTs) are mesenchymal tumors of the gastrointestinal tract that are most commonly found in the stomach. Although GISTs can spread to the liver and peritoneum, metastasis to the skeletal muscle is very rare and only four cases have previously been reported. These cases involved concurrent skeletal metastases of primary GISTs or liver metastases. Here, we report the first case of a distant recurrence in the brachialis muscle after complete remission of an extra-luminal gastric GIST following a wedge resection of the stomach, omental excision, and adjuvant imatinib therapy for one year. Ten months after therapy completion, the patient presented with swelling and tenderness in the left arm. Magnetic resonance imaging revealed a large mass in the brachialis muscle, which showed positivity for c-kit and CD34 upon pathologic examination. This is the first reported case of a solitary distant recurrence of a GIST in the muscle after complete remission had been achieved.

Idiopathic pleuroparenchymal fibroelastosis presenting in recurrent pneumothorax: a case report.

Idiopathic pleuroparenchymal fibroelastosis (PPFE) is a rare, recently classified entity that consists of pleural and subjacent parenchymal fibrosis predominantly in the upper lungs. In an official American Thoracic Society/European Respiratory Society statement in 2013, this disease is introduced as a group of rare idiopathic interstitial pneumonias. We describe a case of a 76-year-old woman with cough and recurrent pneumothorax. She was admitted to our hospital with severe cough at first. High resolution computed tomography (HRCT) disclosed multifocal subpleural consolidations with reticular opacities in both lungs, primarily in the upper lobes, suggesting interstitial pneumonia. Rheumatoid lung was diagnosed initially through an elevated rheumatoid factor, HRCT and surgical biopsy at the right lower lobe. However, one month later, pneumothorax recurred. Surgical biopsy was performed at the right upper lobe at this time. The specimens revealed typical subpleural fibroelastosis. We report this as a first case of idiopathic PPFE in Korea after reviewing the symptoms, imaging and pathologic findings.

Role of high-resolution magnetic resonance imaging in the diagnosis of primary angiitis of the central nervous system.

BACKGROUND: Primary angiitis of the central nervous system (PACNS) is a rare disorder and is often difficult to diagnose due to the lack of a confirmatory test. PACNS can generally be diagnosed based on typical angiographic findings. We describe herein a patient diagnosed with PACNS despite the presence of normal findings on conventional angiography. CASE REPORT: A 44-year-old man with a recent history of ischemic stroke in the right posterior cerebral artery territory developed acute-onset vertigo. Diffusion-weighted imaging revealed an acute infarction within the left posterior inferior cerebellar artery. His medical history was unremarkable except for hyperlipidemia; the initial examination revealed mild gait imbalance. During the 10 days of hospital admission, the patient experienced four recurrent ischemic strokes within the posterior circulation territory (occipital lobe, pons, and cerebellum). He was diagnosed with recurrent cerebral infarctions due to PACNS. The basilar artery exhibited no demonstrable luminal stenosis, but there were direct imaging signs of central nervous system angiitis including wall thickening and contrast enhancement. High-dose intravenous steroid therapy followed by oral prednisolone was administered. There was no further stroke recurrence and follow-up imaging of the arterial walls showed normalization of their characteristics. CONCLUSIONS: The present case emphasizes the importance of wall imaging in the diagnosis and treatment of PACNS.

Brain Gray Matter Deficits in Patients with Chronic Primary Insomnia.

STUDY OBJECTIVE: To investigate the structural changes in patients with chronic primary insomnia and the relationships with clinical features of insomnia. DESIGN: Statistical parametric mapping 8-based voxel-based morphometry was used to identify differences in regional gray and white matter between patients with chronic primary insomnia and normal controls. SETTING: University hospital. PATIENTS AND PARTICIPANTS: Twenty-seven patients and 27 age/sex-matched controls. INTERVENTIONS: Regional differences were compared using two-sample t-tests with age, sex, and intracranial volume as covariates. MEASUREMENTS AND RESULTS: The patients were a mean age of 52.3 y and had a mean history of insomnia of 7.6 y. Patients displayed cognitive deficits in attention, frontal/executive function, and nonverbal memory. Patients also displayed significantly reduced gray matter concentrations (GMCs) in dorsolateral prefrontal and pericentral cortices, superior temporal gyrus, and cerebellum and decreased gray matter volumes in medial frontal and middle temporal gyri compared with control patients with the cluster threshold ≥ 50 voxels at the level of uncorrected P < 0.001. Negative correlations were found between GMC of the prefrontal cortex and insomnia severity and the wakefulness after sleep onset, and between GMC of pericentral cortex and sleep latencies. None of the findings continued to be significant after correction for multiple comparisons. CONCLUSIONS: We found gray matter deficits in multiple brain regions including bilateral frontal lobes in patients with psychophysiologic insomnia. Gray matter deficit of the pericentral and lateral temporal areas may be associated with the difficulties in sleep initiation and maintenance. It is still unclear whether gray matter reductions are a preexisting abnormality or a consequence of insomnia. CITATION: Joo EY; Noh HJ; Kim JS; Koo DL; Kim D; Hwang KJ; Kim JY; Kim ST; Kim MR; Hong SB. Brain gray matter deficits in patients with chronic primary insomnia. SLEEP 2013;36(7):999-1007.