mTOR signalling controls the formation of smooth muscle cell-derived luminal myofibroblasts during vasculitis.

The accumulation of myofibroblasts within the intimal layer of inflamed blood vessels is a potentially catastrophic complication of vasculitis, which can lead to arterial stenosis and ischaemia. In this study, we have investigated how these luminal myofibroblasts develop during Kawasaki disease (KD), a paediatric vasculitis typically involving the coronary arteries. By performing lineage tracing studies in a murine model of KD, we reveal that luminal myofibroblasts develop independently of adventitial fibroblasts and endothelial cells, and instead derive from smooth muscle cells (SMCs). Notably, the emergence of SMC-derived luminal myofibroblasts-in both mice and patients with KD, Takayasu's arteritis and Giant Cell arteritis-coincided with activation of the mechanistic target of rapamycin (mTOR) signalling pathway. Moreover, SMC-specific deletion of mTOR signalling, or pharmacological inhibition, abrogated the emergence of luminal myofibroblasts. Thus, mTOR is an intrinsic and essential regulator of luminal myofibroblast formation that is activated in vasculitis patients and therapeutically tractable. These findings provide molecular insight into the pathogenesis of coronary artery stenosis and identify mTOR as a therapeutic target in vasculitis.

A Call to Action to Improve Cardiac Arrest Outcomes: A Report From the National Summit for Cardiac Arrest.

Sudden cardiac arrest (SCA) represents a major cause of premature mortality globally, with enormous impact and financial cost to victims, families, and communities. SCA prevention should be considered a health priority in Australia. National Cardiac Arrest Summits were held in June 2022 and March 2023, with inclusion from multi-faceted endeavours related to SCA prevention. It was agreed to establish a multidisciplinary Australian Sudden Cardiac Arrest Alliance (AuSCAA) working group charged with developing a national unified strategy, with clear and measurable quality indicators and standardised outcome measures, to amplify the goal of SCA prevention throughout Australia. A multi-faceted prevention strategy will include i) endeavours to progress community awareness, ii) improved fundamental mechanistic understanding, iii) implementation of best-practice resuscitation strategies for all demographics and locations, iv) secondary risk assessment directed to family members, and v) development of (near) real-time registry of cardiac arrest cases to inform areas of need and effectiveness of interventions. Together, we can and should reduce the impact of SCA in Australia.

A retrospective review of methylamphetamine detected in child deaths reported to the Victorian Coroner, Australia.

This study investigated methylamphetamine (MA) exposures in the deaths of children (≤ 12 years old) reported to the Coroner in the state of Victoria, Australia, between 2011 and 2020. Demographics, autopsy findings including the cause of death, self-reported prenatal or caregiver drug use, child protection services information, and toxicological findings were summarized by descriptive statistics. Validated methods of liquid chromatography-tandem mass spectrometry were used in the analysis of drugs. There were 50 child deaths with MA detected in blood, urine, and/or hair with 64% (n = 32) identified in 2018-2020. Most children were 1-365 days old (66%, n = 33) and the cause of death was unascertained in 62% (n = 31) of cases. MA was toxicologically confirmed in hair (94%, n = 47) significantly more than blood (18%, n = 9). Prenatal or caregiver drug use was self-reported in 44% (n = 22) and 42% (n = 21) of cases, respectively. Moreover, only 54% (n = 27) of deceased children were a child protection client at their time of death. These findings suggest the number of deceased children exposed to MA has increased over the past 10 years, which is consistent with the greater supply of crystal MA in the Australian community. Hair analysis provided additional means to identify cases that were unknown to child protection services and may have implications for other children in the same drug exposure environment.

Inconsistent discharge diagnoses for young cardiac arrest episodes: insights from a statewide registry.

BACKGROUND: Administrative coding of out-of-hospital cardiac arrest (OHCA) is heterogeneous, with the prevalence of noninformative diagnoses uncertain. AIM: To characterize the prevalence and type of non-informative diagnoses in a young cardiac arrest population. METHODS: Hospital discharge diagnoses provided to a statewide OHCA registry were characterised as either 'informative' or 'noninformative.' Informative diagnoses stated an OHCA had occurred or defined OHCA as occurring due to coronary artery disease, cardiomyopathy, channelopathy, definite noncardiac cause, or no known cause. Noninformative diagnoses were blank, stated presenting cardiac rhythm only, provided irrelevant information or presented a complication of the OHCA as the main diagnosis. Characteristics of patients receiving informative versus noninformative diagnoses were compared. RESULTS: Of 1479 patients with OHCA aged 1 to 50 years, 290 patients were admitted to 15 hospitals. Ninety diagnoses (31.0%) were noninformative (arrest rhythm = 50, blank = 21, complication = 10 and irrelevant = 9). Two hundred diagnoses (69.0%) were informative (cardiac arrest = 84, coronary artery disease = 54, noncardiac diagnosis = 48, cardiomyopathy = 8, arrhythmia disorder = 4 and unascertained = 2). Only 10 diagnoses (3.5%) included both OHCA and an underlying cause. Patients receiving a noninformative diagnosis were more likely to have survived OHCA or been referred for forensic assessment (P = 0.011) and had longer median length of stay (9 vs 5 days, P = 0.0019). CONCLUSION: Almost one third of diagnoses for young patients discharged after an OHCA included neither OHCA nor any underlying cause. Underestimating the burden of OHCA impacts ongoing patient and at-risk family care, data sampling strategies, international statistics and research funding.

Histiocytoid cardiomyopathy presenting as sudden death in an 18-month-old infant.

Histiocytoid cardiomyopathy (HC) is an arrhythmogenic disorder, usually involving children under two years of age with a strong Caucasian and female predominance. The disease is fatal in the vast majority and diagnosis is nearly always established at autopsy, but this is only possible with adequate myocardial sampling. Meticulous gross and histological examination of the heart in collaboration with a cardiovascular-trained pathologist maximises the opportunity to make specific diagnoses (and therefore rule out the differentials of SIDS, SUDC and child abuse), guide genetic testing, and inform potentially life-saving medical interventions for blood relations. We present a typical HC case presenting as sudden death, without prodrome, in a previously healthy 18-month-old boy. The disease is characterised histologically by discrete groups of enlarged, polygonal histiocyte-like cells with distinct margins and abundant faintly eosinophilic foamy cytoplasm. Cells often contain coarse granules, microvacuoles and irregular, round nuclei. In our case, dysplastic fascicles were predominantly located immediately deep to the endocardium of the left ventricle. We report our own autopsy findings with histological images, and discuss the expected clinical, morphological and ultrastructural features of the disease.

Fatal gastric volvulus: forensic pathology considerations and postmortem CT findings.

Gastric volvulus is a rare cause of gastric obstruction, due to the rotation of the stomach by more than 180°. It is a rare but life-threatening medical emergency that is considered difficult to diagnose at the initial clinical presentation. Forensic pathologists may be presented with gastric volvulus in several ways, for instance, as a cause of sudden and unexpected death or in the context of suspected clinical errors. The post-mortem examination of gastric volvulus may be challenging, due to the specific technical issues it presents and the various mechanisms by which volvulus may cause death. We therefore present five cases of gastric volvulus that in combination represent almost the entire spectrum of presentations and post-mortem findings, to discuss how gastric volvulus may come to the attention of a forensic pathologist, the approach and findings at post-mortem examination (including post-mortem CT), and the variety of mechanisms by which gastric volvulus may result in death.

Causes, circumstances, and potential preventability of cardiac arrest in the young: insights from a state-wide clinical and forensic registry.

AIMS: The causes, circumstances, and preventability of young sudden cardiac arrest remain uncertain. METHODS AND RESULTS: A prospective state-wide multi-source registry identified all out-of-hospital cardiac arrests (OHCAs) in 1-50 year olds in Victoria, Australia, from 2019 to 2021. Cases were adjudicated using hospital and forensic records, clinic assessments and interviews of survivors and family members. For confirmed cardiac causes of OHCA, circumstances and cardiac history were collected. National time-use data was used to contextualize circumstances. 1319 OHCAs were included. 725 (55.0%) cases had a cardiac aetiology of OHCA, with coronary disease (n = 314, 23.8%) the most common pathology. Drug toxicity (n = 226, 17.1%) was the most common non-cardiac cause of OHCA and the second-most common cause overall. OHCAs were most likely to occur in sleep (n = 233, 41.2%). However, when compared to the typical Australian day, OHCAs occurred disproportionately more commonly during exercise (9% of patients vs. 1.3% of typical day, P = 0.018) and less commonly while sedentary (39.6 vs. 54.6%, P = 0.047). 38.2% of patients had known standard modifiable cardiovascular risk factors. 77% of patients with a cardiac cause of OHCA had not reported cardiac symptoms nor been evaluated by a cardiologist prior to their OHCA. CONCLUSION: Approximately half of OHCAs in the young have a cardiac cause, with coronary disease and drug toxicity dominant aetiologies. OHCAs disproportionately occur during exercise. Of patients with cardiac cause of OHCA, almost two-thirds have no standard modifiable cardiovascular risk factors, and more than three-quarters had no prior warning symptoms or interaction with a cardiologist.

Effect of plane of nutrition and analgesic drug treatment on wound healing and pain following cautery disbudding in preweaning dairy calves.

The objective of this study was to determine the effect of a biologically normal plane of nutrition compared with a limited plane on the primary outcome wound healing, and one dose of nonsteroidal anti-inflammatory drug (NSAID) compared with 2 on the secondary outcomes: lying behavior, haptoglobin concentrations, and mechanical nociceptive threshold (MNT) in calves disbudded via cautery iron. Eighty female Holstein calves were enrolled at birth, individually housed, and fed via a Calf Rail system (Förster Technik). A 2 × 2 factorial design was used to assess the effect of plane of nutrition and an additional NSAID. Calves were randomly assigned to a biologically normal plane of nutrition (BN; offered up to 15 L/d) or a limited plane (LP; offered up to 6 L/d) and to receive one or 2 doses of meloxicam. All calves received a lidocaine cornual nerve block and a subcutaneous injection of meloxicam 15 min before cautery disbudding at 18 to 25 d of age, and half the calves received an additional injection of meloxicam (0.5 mg/kg) 3 d after disbudding. Tissue type present, wound diameter, and wound depth were evaluated 2 times per week for 7 to 8 wk as measures of wound healing, lying behavior was recorded beginning 1 to 2 wk before disbudding until 7 to 8 wk after as a behavioral indicator of pain, haptoglobin concentrations were measured once per day for 7 d after disbudding, and MNT was evaluated 2 times/wk for 3 wk. Survival analyses were analyzed using Cox regression models (wound healing) and continuous data were analyzed using mixed-effect linear regression models. Only 12% of horn buds were completely healed by 7 to 8 wk after disbudding and 54% had re-epithelized at this time. At any time, wounds from BN calves were more likely to have had re-epithelization occur compared with wounds from LP calves (hazard ratio: 1.93, 95% CI: 1.18-3.14). Wounds from calves that received only one dose of NSAID were more likely to have re-epithelization occur, compared with wounds from calves given 2 doses (hazard ratio: 1.87, 95% CI: 1.15-3.05). Wounds from BN calves had smaller diameters and depths over time beginning on wk 3 compared with LP calves. Wounds from calves that received an additional NSAID had larger diameters and depths over time beginning on wk 4 and 3 respectively, compared with calves that only received one dose of NSAID. Calves that received an extra NSAID tended to be less sensitive 7, 10, and 17 d after disbudding compared with calves that only received one dose and spent less time lying in the week after disbudding. Calves on the BN milk program were more active compared with LP calves with lower lying times, fewer lying bouts per day, and longer average lying bouts. Our results indicate that a BN milk feeding program for calves can result in faster healing times and more activity, and that providing an extra NSAID 3 d after disbudding appears to slow the healing process but may result in less pain experienced by the calf 1 to 2 wk after the procedure. This study is also among the first to demonstrate that after the complete removal of the horn bud, wounds can take more than 8 weeks to re-epithelize and fully heal.

Higher rates but similar causes of young out-of-hospital cardiac arrest in rural Australian patients.

OBJECTIVE: To determine whether young rural Australians have higher rates or different underlying causes of out-of-hospital cardiac arrest (OHCA). DESIGN: A case-control design identified patients experiencing an OHCA, then compared annual OHCA rates and underlying causes in rural versus metropolitan Victoria. OHCA causes were defined as either cardiac or non-cardiac, with specific aetiologies including coronary disease, cardiomyopathy, unascertained cause of arrest, drug toxicity, respiratory event, neurological event and other cardiac and non-cardiac. For OHCAs with confirmed cardiac aetiology, cardiovascular risk profiles were compared. SETTING: A state-wide prospective OHCA registry (combining ambulance, hospital and forensic data) in the state of Victoria, Australia (population 6.5 million). PARTICIPANTS: Victorians aged 1-50 years old experienced an OHCA between April 2019 and April 2020. MAIN OUTCOME MEASURES: Rates and underlying causes of OHCA in young rural and metropolitan Victorians. RESULTS: Rates of young OHCA were higher in rural areas (OHCA 22.5 per 100 000 rural residents vs. 13.4 per 100 000 metropolitan residents, standardised incidence ratio 168 (95% CI 101-235); confirmed cardiac cause of arrest 12.1 per 100 000 rural residents versus 7.5 per 100 000 metropolitan residents, standardised incidence ratio 161 (95% CI 71-251). The underlying causation of the OHCA and cardiovascular risk factor burden did not differ between rural and metropolitan areas. CONCLUSION: Higher rates of OHCA occur in young rural patients, with standardised incidence ratio of 168 compared to young metropolitan residents. Rural status did not influence causes of cardiac arrest or known cardiovascular risk factor burden in young patients experiencing OHCA.

Multisystem Inflammatory Syndrome in Children: Survey of Protocols for Early Hospital Evaluation and Management.

OBJECTIVE: To describe the similarities and differences in the evaluation and treatment of multisystem inflammatory syndrome in children (MIS-C) at hospitals in the US. STUDY DESIGN: We conducted a cross-sectional survey from June 16 to July 16, 2020, of US children's hospitals regarding protocols for management of patients with MIS-C. Elements included characteristics of the hospital, clinical definition of MIS-C, evaluation, treatment, and follow-up. We summarized key findings and compared results from centers in which >5 patients had been treated vs those in which ≤5 patients had been treated. RESULTS: In all, 40 centers of varying size and experience with MIS-C participated in this protocol survey. Overall, 21 of 40 centers required only 1 day of fever for MIS-C to be considered. In the evaluation of patients, there was often a tiered approach. Intravenous immunoglobulin was the most widely recommended medication to treat MIS-C (98% of centers). Corticosteroids were listed in 93% of protocols primarily for moderate or severe cases. Aspirin was commonly recommended for mild cases, whereas heparin or low molecular weight heparin were to be used primarily in severe cases. In severe cases, anakinra and vasopressors frequently were recommended; 39 of 40 centers recommended follow-up with cardiology. There were similar findings between centers in which >5 patients vs ≤5 patients had been managed. Supplemental materials containing hospital protocols are provided. CONCLUSIONS: There are many similarities yet key differences between hospital protocols for MIS-C. These findings can help healthcare providers learn from others regarding options for managing MIS-C.

The End Unexplained Cardiac Death (EndUCD) Registry for Young Australian Sudden Cardiac Arrest.

In 2019, the first multi-source registry of sudden cardiac arrest and death for patients aged 1-50 years launched in Victoria, Australia. Sudden cardiac arrest (SCA) affects approximately fifteen hundred younger Victorians per year. The End Unexplained Cardiac Death (EndUCD) Registry enrols SCA/death (D) cases aged 1-50 years, providing family screening, access to psychological support through clinical sites and creating a genetic biorepository for whole-genome sequencing. The registry will support clear pathways of cardiac assessment, epidemiological profiling and routine family screening and psychological support.

Can post-mortem coronary artery calcium scores aid diagnosis in young sudden death?

This study sought to explore the feasibility and utility of post-mortem coronary artery calcium (CAC) scoring in identifying patients with ischemic heart disease as cause of sudden death. 100 deceased patients aged 18-50 years underwent post-mortem examination in the setting of sudden death. At post-mortem, fifty cases were determined to have ischemic heart disease, and fifty had death attributed to trauma or unascertained causes. The CAC score was calculated in a blinded manner from post-mortem CTs performed on all cases. CAC scores were assessable in 97 non-decomposed cases (feasibility 97%). The median CAC score was 88 Agatston units [IQR 0-286] in patients deceased from ischemic heart disease vs 0 [IQR 0-0] in patients deceased from other causes (p < 0.0001). Presence of any coronary calcification differed significantly between ischemic heart disease and non-ischemic groups (adjusted odds ratio 10.7, 95% CI 3.2-35.5). All cases with a CAC score > 100 (n = 22) had ischemic heart disease as the cause of death. Fifteen cases had a CAC score of zero but severe coronary disease at post-mortem examination. Post-mortem CAC scoring is highly feasible. An elevated CAC score in cases 18-50 years old with sudden death predicts ischemic heart disease at post-mortem examination. However, a CAC score of zero does not exclude significant coronary artery disease. Post-mortem CAC score may be considered as a further assessment tool to help predict likely cause of death when there is an objection to or unavailability of post-mortem examination.

Insights into sudden cardiac death: exploring the potential relevance of non-diagnostic autopsy findings.

AIMS: Unexplained sudden cardiac death (SCD) may be attributable to cardiogenetic disease. Presence or absence of autopsy anomalies detected following premature sudden death direct appropriate clinical evaluation of at-risk relatives towards inherited cardiomyopathies or primary arrhythmia syndromes, respectively. We investigated the relevance of non-diagnostic pathological abnormalities of indeterminate causality (uncertain) such as myocardial hypertrophy, fibrosis, or inflammatory infiltrates to SCD. METHODS AND RESULTS: At-risk relatives of unexplained SCD cases aged 1-64 years without prior cardiac disease (n = 98) with either normal and negative (40%, true sudden arrhythmic death syndrome; SADS) or isolated non-diagnostic (60%, uncertain sudden unexplained death; SUD) cardiac histological autopsy findings at a central forensic pathology unit were referred to the regional unexplained SCD clinic for clinical cardiac phenotyping. Uncertain SUD were older than true SADS cases (31.8 years vs. 21.1 years, P < 0.001). A cardiogenetic diagnosis was established in 24 families (24.5%) following investigation of 346 referred relatives. The proportions of uncertain SUD and true SADS explained by familial cardiogenetic diagnoses were similar (20% vs. 31%, P = 0.34, respectively), with primary arrhythmia syndromes predominating. Unexplained SCD cases were more likely than matched non-cardiac premature death controls to demonstrate at least one uncertain autopsy finding (P < 0.001). CONCLUSION: Primary arrhythmia syndromes predominate as familial cardiogenetic diagnoses amongst both uncertain SUD and true SADS cases. Non-diagnostic or uncertain histological findings associate with SUD, though cannot be attributed a causative status. At-risk relatives of uncertain SUD cases should be evaluated for phenotypic evidence of both ion channel disorders and cardiomyopathies.

P190RhoGAP prevents mitotic spindle fragmentation and is required to activate Aurora A kinase at acentriolar poles.

Assembly of the mitotic spindle is essential for proper chromosome segregation during mitosis. Maintenance of spindle poles requires precise regulation of kinesin- and dynein-generated forces, and improper regulation of these forces disrupts pole integrity leading to pole fragmentation. The formation and function of the mitotic spindle are regulated by many proteins, including Aurora A kinase and the motor proteins Kif2a and Eg5. Here, we characterize a surprising role for the RhoA GTPase-activating protein, p190RhoGAP, in regulating the mitotic spindle. We show that cells depleted of p190RhoGAP arrest for long periods in mitosis during which cells go through multiple transitions between having bipolar and multipolar spindles. Most of the p190RhoGAP-depleted cells finally achieve a stable bipolar attachment and proceed through anaphase. The multipolar spindle phenotype can be rescued by low doses of an Eg5 inhibitor. Moreover, we show that p190RhoGAP-depleted multipolar cells localize Aurora A to all the poles, but the kinase is only activated at the two centriolar poles. Overall, our data identify an unappreciated connection between p190RhoGAP and the proteins that control spindle poles including Aurora A kinase and Eg5 that is required to prevent or correct spindle pole fragmentation.

Toxoplasma gondii induces FAK-Src-STAT3 signaling during infection of host cells that prevents parasite targeting by autophagy.

Targeting of Toxoplasma gondii by autophagy is an effective mechanism by which host cells kill the protozoan. Thus, the parasite must avoid autophagic targeting to survive. Here we show that the mammalian cytoplasmic molecule Focal Adhesion Kinase (FAK) becomes activated during invasion of host cells. Activated FAK appears to accompany the formation of the moving junction (as assessed by expression the parasite protein RON4). FAK activation was inhibited by approaches that impaired ß1 and ß3 integrin signaling. FAK caused activation of Src that in turn mediated Epidermal Growth Factor Receptor (EGFR) phosphorylation at the unique Y845 residue. Expression of Src-resistant Y845F EGFR mutant markedly inhibited ROP16-independent activation of STAT3 in host cells. Activation of FAK, Y845 EGFR or STAT3 prevented activation of PKR and eIF2α, key stimulators of autophagy. Genetic or pharmacologic inhibition of FAK, Src, EGFR phosphorylation at Y845, or STAT3 caused accumulation of the autophagy protein LC3 and LAMP-1 around the parasite and parasite killing dependent on autophagy proteins (ULK1 and Beclin 1) and lysosomal enzymes. Parasite killing was inhibited by expression of dominant negative PKR. Thus, T. gondii activates a FAK→Src→Y845-EGFR→STAT3 signaling axis within mammalian cells, thereby enabling the parasite to survive by avoiding autophagic targeting through a mechanism likely dependent on preventing activation of PKR and eIF2α.

A complex of p190RhoGAP-A and anillin modulates RhoA-GTP and the cytokinetic furrow in human cells.

The cytokinetic furrow is organized by the RhoA GTPase, which recruits actin and myosin II to the furrow and drives contractility. Here, we show that the RhoA GTPase-activting protein (GAP) p190RhoGAP-A (also known as ARHGAP35) has a role in cytokinesis and is involved in regulating levels of RhoA-GTP and contractility. Cells depleted of p190RhoGAP-A accumulate high levels of RhoA-GTP and markers of high RhoA activity in the furrow, resulting in failure of the cytokinetic furrow to progress to abscission. The loss of p190RhoGAP-A can be rescued by a low dose of the myosin II inhibitor blebbistatin, suggesting that cells fail cytokinesis because they have too much myosin activity. p190RhoGAP-A binds the cytokinetic organizer anillin, and mutants of p190RhoGAP-A that are unable to bind anillin or unable to inactivate RhoA fail to rescue cytokinesis defects in p190RhoGAP-A-depleted cells. Taken together, these data demonstrate that a complex of p190RhoGAP-A and anillin modulates RhoA-GTP levels in the cytokinetic furrow to ensure progression of cytokinesis.

The effect of post-mortem computed tomography angiography (PMCTA) using water-soluble, iodine-based radiographic contrast on histological analysis of the liver, kidneys and left ventricle of the heart.

The purpose of this study was to investigate the impact of post-mortem computed-tomography angiography (PMCTA) on the histology of the liver, kidneys and heart. Multiple tissue cores were collected from the liver, left and right kidneys and left ventricle utilizing CT-guided biopsy. Subsequent whole body PMCTA was performed using a solution of polyethylene glycol and iodinated radiographic contrast, and an embalming pump. Corresponding biopsy cores were collected at autopsy, and blinded histology analysis assessing for PMCTA-induced histology artefact was performed. The blinded analysis of pre-PMCTA and post-PMCTA biopsy samples demonstrated that whole body PMCTA had no effect on the histological analyses of the liver (0%, CI = 0-13.7%), left ventricle of the heart (0%, CI = 0-36.9%) and right kidney (0%, CI = 13.2%), however likely caused increased Bowman's capsule spaces in the left kidney of one case (4%, CI = 0.01-20.4%). Other artefactual histological changes identified included eosinophilic material in the liver, whiter interstitium and dilated tubules in kidney samples, and autolysis-related changes, however these could not be categorically attributed to the PMCTA procedure. PMCTA causes zero or minimal effect to the histological examination of the liver, left kidney, right kidney and left ventricle, and as such performing PMCTA prior to autopsy is unlikely to impact autopsy histological results in these organs.

Sudden cardiac death while playing Australian Rules football: a retrospective 14 year review.

Australian Rules football is a sport which evolved from Gaelic football and which is played by a large number of predominantly male participants in a number of countries. The highest participation rates are in the southern states of Australia. A retrospective review over a period of 14 years identified 14 cases of sudden cardiac death that occurred in individuals while playing the sport. All were male and ranged in age from 13 to 36 years with a mean and median age of 23 years. A spectrum of cardiac causes was identified including coronary artery atherosclerosis, myocarditis, anomalous coronary artery anatomy, arrhythmogenic right ventricular cardiomyopathy, and healed Kawasaki disease. In 5 cases the heart was morphologically normal raising the possibility of a channelopathy. No traumatic deaths were identified. Some of the individuals had experienced symptoms prior to the fatal episode and the role of pre participation screening in reducing mortality is discussed.

The human papillomavirus E7 proteins associate with p190RhoGAP and alter its function.

UNLABELLED: Using mass spectrometry, we identified p190RhoGAP (p190) as a binding partner of human papillomavirus 16 (HPV16) E7. p190 belongs to the GTPase activating protein (GAP) family and is one of the primary GAPs for RhoA. GAPs stimulate the intrinsic GTPase activity of the Rho proteins, leading to Rho inactivation and influencing numerous biological processes. RhoA is one of the best-characterized Rho proteins and is specifically involved in formation of focal adhesions and stress fibers, thereby regulating cell migration and cell spreading. Since this is the first report that E7 associates with p190, we carried out detailed interaction studies. We show that E7 proteins from other HPV types also bind p190. Furthermore, we found that conserved region 3 (CR3) of E7 and the middle domain of p190 are important for this interaction. More specifically, we identified two residues in CR3 of E7 that are necessary for p190 binding and used mutants of E7 with mutations of these residues to determine the biological consequences of the E7-p190 interaction. Our data suggest that the interaction of E7 with p190 dysregulates this GAP and alters the actin cytoskeleton. We also found that this interaction negatively regulates cell spreading on a fibronectin substrate and therefore likely contributes to important aspects of the HPV life cycle or HPV-induced tumorigenesis. IMPORTANCE: This study identifies p190RhoGAP as a novel cellular binding partner for the human papillomavirus (HPV) E7 protein. Our study shows that a large number of different HPV E7 proteins bind p190RhoGAP, and it identifies regions in both E7 and p190RhoGAP which are important for the interaction to occur. This study also highlights the likelihood that the E7-p190RhoGAP interaction may have important biological consequences related to actin organization in the infected cell. These changes could be an important contributor to the viral life cycle and during progression to cancer in HPV-infected cells. Importantly, this work also emphasizes the need for further study in a field which has largely been unexplored as it relates to the HPV life cycle and HPV-induced transformation.

Epidermal growth factor-receptor activation modulates Src-dependent resistance to lapatinib in breast cancer models.

INTRODUCTION: Src tyrosine kinase overactivation has been correlated with a poor response to human epidermal growth factor receptor 2 (HER2) inhibitors in breast cancer. To identify the mechanism by which Src overexpression sustains this resistance, we tested a panel of breast cancer cell lines either sensitive or resistant to lapatinib. METHODS: To determine the role of Src in lapatinib resistance, we evaluated the effects of Src inhibition/silencing in vitro on survival, migration, and invasion of lapatinib-resistant cells. In vivo experiments were performed in JIMT-1 lapatinib-resistant cells orthotopically implanted in nude mice. We used artificial metastasis assays to evaluate the effect of Src inhibition on the invasiveness of lapatinib-resistant cells. Src-dependent signal transduction was investigated with Western blot and ELISA analyses. RESULTS: Src activation was higher in lapatinib-resistant than in lapatinib-sensitive cells. The selective small-molecule Src inhibitor saracatinib combined with lapatinib synergistically inhibited the proliferation, migration, and invasion of lapatinib-resistant cells. Saracatinib combined with lapatinib significantly prolonged survival of JIMT-1-xenografted mice compared with saracatinib alone, and impaired the formation of lung metastases. Unexpectedly, in lapatinib-resistant cells, Src preferentially interacted with epidermal growth factor receptor (EGFR) rather than with HER2. Moreover, EGFR targeting and lapatinib synergistically inhibited survival, migration, and invasion of resistant cells, thereby counteracting Src-mediated resistance. These findings demonstrate that Src activation in lapatinib-resistant cells depends on EGFR-dependent rather than on HER2-dependent signaling. CONCLUSIONS: Complete pharmacologic EGFR/HER2 inhibition is required to reverse Src-dependent resistance to lapatinib in breast cancer.