Receptor tyrosine kinases and TLR/IL1Rs unexpectedly activate myeloid cell PI3kγ, a single convergent point promoting tumor inflammation and progression

We show that imatinib, nilotinib, and dasatinib possess weak off-target activity against RAF and, therefore, drive paradoxical activation of BRAF and CRAF in a RAS-dependent manner. Critically, because RAS is activated by BCR-ABL, in drug-resistant chronic myeloid leukemia (CML) cells, RAS activity persists in the presence of these drugs, driving paradoxical activation of BRAF, CRAF, MEK, and ERK, and leading to an unexpected dependency on the pathway. Consequently, nilotinib synergizes with MEK inhibitors to kill drug-resistant CML cells and block tumor growth in mice. Thus, we show that imatinib, nilotinib, and dasatinib drive paradoxical RAF/MEK/ERK pathway activation and have uncovered a synthetic lethal interaction that can be used to kill drug-resistant CML cells in vitro and in vivo.     

Aberrant right coronary artery origin from the left aortic sinus with interarterial course manifesting acute myocardial infarction

A 43-year-old patient was admitted to hospital because of an inferior-posterior myocardial infarction. The admission electrocardiogram was suggestive of a right coronary artery (RCA) culprit lesion. Coronary angiography following successful thrombolysis revealed a normal left system and mild intraluminal disease of the dominant RCA, which arose from the left aortic sinus and travelled an interarterial course; the latter was depicted in a subsequent computed tomographic angiogram. The lack of ST segment elevation in V4R and the absence of right ventricular wall motion abnormalities on echocardiography precluded the proximal ectopic vessel from being the culprit. The patient was managed medically; one year following discharge, he is asymptomatic. In cases of aberrant anomalous origin of a coronary artery from the opposite sinus with interarterial course, the proximal ectopic vessel is intussuscepted within the aortic wall, potentially leading to ischemia. The present article highlights that, although medical treatment in cases of such an aberrant RCA without apparent ischemia-driven sequelae may be valid, the need for interventional treatment could be substantiated following investigation of the anatomofunctional features of the intussuscepted proximal ectopic segment with intravascular ultrasound.     

Anomalous origin of right coronary artery from the mid-left anterior descending artery

We report a case of a right coronary artery (RCA) originating from the mid-left anterior descending artery (LAD) coursing around and in front of the pulmonary artery. The anomaly was discovered incidentally during cardiac catheterization. It is only the second case of such an anomaly ever described.     

Catheter closure of atrial septal defects using the Cocoon septal occluder: Preliminary results of a European multicenter study

Despite its simplicity, device closure of atrial septal defects is still associated with rare but potentially lethal complications. In this prospective non-randomized multicenter study we investigated the safety and efficacy of the Cocoon septal occluder (CSO) for closure of atrial septal defects (ASDs) in 92 patients. Median age of the patients was 10.5 years (range 3–61 years) and median weight was 25 kg (range 13–65 kg). The device is an improved new generation double disc design made of Nitinol wire mesh that is coated with platinum using NanoFusion technology. The discs are connected by a waist with diameter ranging from 6 mm to 40 mm with 2 mm increments. All patients completed a 3-month follow-up. Mean ASD diameter was 21 ± 7 mm (range 10–35 mm), while the mean device diameter was 24 ± 8 mm (range 14–40 mm). The CSO was permanently implanted in all 92 patients. Complete echocardiographic closure of the defect immediately after the procedure or at the one month follow-up, was observed in all 92 patients (100%). No device-related complications were observed during the procedure or at short-term follow-up (range 3–12 months). Our preliminary results indicate that CSO is a promising device for transcatheter closure of ASDs. Further studies are required to document its efficacy, safety and long-term results in a larger patient population.     

Isolation of esophageal stem cells with potential for therapy

Purpose

Long-gap esophageal atresia represents a significant challenge for pediatric surgeons and current surgical approaches are associated with significant morbidity. A tissue-engineered esophagus, comprising cells seeded onto a scaffold, represents a therapeutic alternative. In this study, we aimed to determine the optimal techniques for isolation and culture of mouse esophageal epithelial cells and to isolate CD34-positive esophageal epithelial stem cells from cadaveric mouse specimens.

Methods

Primary epithelial cells were isolated from mouse esophagi by enzymatic dissociation from the mucosal layer (Dispase, Trypsin/EDTA) using three different protocols. In protocol A, isolated mucosa was minced and incubated with trypsin once. In protocol B, intact mucosal sheets underwent two trypsin incubations yielding a single-cell suspension. In protocol C, intact mucosa explants were plated epithelial side down. Epithelial cells were cultured on collagen-coated wells.

Results

Initial findings showed that Protocol B gave the best results in terms of yield, viability, and least contamination with different cell types and microbes. Esophageal epithelial cells isolated using Protocol B were stained for CD34 and sorted using fluorescence-activated cell sorting (FACS). Of the total cells sorted, 8.3 % (2–11.3) [%median (range)] were CD34 positive.

Conclusions

Our results demonstrate that mouse esophageal epithelial cells can be successfully isolated from fresh mouse esophagi using two consecutive trypsin incubations of intact mucosal sheets. Furthermore, the cells obtained using this method were successfully stained for CD34, a putative esophageal epithelial stem cell marker. Further research into the factors necessary for the successful proliferation of CD34 positive stem cell lines is needed to progress toward clinical application.     

Conservation of endemic and threatened wildlife: molecular forensic DNA against poaching of the Cypriot mouflon (Ovis orientalis ophion, Bovidae)

Molecular DNA techniques in combination with appropriate reference population database and statistical methods are fundamental tools to forensic wildlife investigations. This is even more relevant when taxa with uncertain systematics are involved, as is the case of the genus Ovis (Bovidae), whose evolution has been influenced by multiple events of domestication. The Cypriot mouflon, Ovis orientalis ophion, a protected subspecies endemic to Cyprus, is threatened by poaching. This study deals with a case of alleged poaching that occurred in Cyprus (September, 2010). A car did not stop at a checkpoint and when finally blocked by the police, several bloodstained exhibits (n=12) were recovered. Three recently deceased mouflons were found by game wardens at the roadside. The Cyprus Veterinary Services established that these animals had been killed by gunshot. As part of the investigation, DNA testing was performed to establish if there was a link between the dead mouflons and the bloodstained exhibits. The mitochondrial Cytochrome-b gene (Cyt-b) and 12 loci of microsatellite DNA were used as markers. The Cyt-b sequences were obtained from 11 exhibits. They were the same as each other and the same as the single haplotype obtained from the three dead mouflons and all the investigated wild Cypriot mouflons (20 individuals). A database of wild mouflons (47 individuals) from which the unknown samples may have originated was generated. The probability of identity (P(ID)) of the microsatellite panel, computed by genotyping all 47 wild mouflons (10 selected loci, P(ID)=10(-5)), allowed us to assign nine exhibits to two out of the three carcasses (seven with very strong support: Likelihood Ratio, LR>3000 and Random Match Probability, RMP, <10(-3)). This study represents the first genetic reference for the Cypriot mouflon and the first published material of forensic wildlife investigations in Cyprus.     

Syncope in a Patient With Giant Left Main Coronary Aneurysm: Is There a Link With Ventricular Arrhythmias?

Giant aneurysm of the left main coronary artery is exceedingly rare and accounts for less than 2% of patients undergoing coronary angiography. The etiology varies depending on the patient''s age and geographic area, but half are of atherosclerotic origin. In most cases, coronary aneurysms are asymptomatic, however, symptomatic patients present with symptoms characteristic of coronary artery disease such as chest pain (angina pectoris), myocardial infarction, congestive heart failure, and even sudden death. Coronary angiography is considered the gold standard tool to determine the presence or absence of coronary artery disease, and if present, its size and location. Herein, we report a case of giant aneurysm of the left main coronary artery presenting as syncope and documented nonsustained ventricular tachycardia.     

Evidence-based management of pregnant women with sickle cell disease in high-income countries

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Isolated single coronary artery with dual right coronary artery distribution

We present the case of a 76-year-old patient in whom coronary angiography, performed due to non-ST-segment elevation myocardial infarction, revealed an isolated single coronary (SCA) artery with dual right coronary artery (RCA) distribution. One RCA arose from the mid segment of the left anterior descending (LAD) artery and followed a prepulmonic course to the right, while the other RCA arose as the terminal extension of the left circumflex artery beyond the crux cordis. This is the second reported case of the combination of these two variants of SCA and the first such case in which the LAD-derived RCA originated as a single branch. Furthermore, this is the first report presenting a sinus node artery with origin from an ectopic LAD-connected RCA. The clinical implications of this rare coronary artery pattern are discussed.     

PI3Kα activates integrin α4β1 to establish a metastatic niche in lymph nodes

Lymph nodes are initial sites of tumor metastasis, yet whether the lymph node microenvironment actively promotes tumor metastasis remains unknown. We show here that VEGF-C/PI3Kα-driven remodeling of lymph nodes promotes tumor metastasis by activating integrin α4β1 on lymph node lymphatic endothelium. Activated integrin α4β1 promotes expansion of the lymphatic endothelium in lymph nodes and serves as an adhesive ligand that captures vascular cell adhesion molecule 1 (VCAM-1)⁺ metastatic tumor cells, thereby promoting lymph node metastasis. Experimental induction of α4β1 expression in lymph nodes is sufficient to promote tumor cell adhesion to lymphatic endothelium and lymph node metastasis in vivo, whereas genetic or pharmacological blockade of integrin α4β1 or VCAM-1 inhibits it. As lymph node metastases accurately predict poor disease outcome, and integrin α4β1 is a biomarker of lymphatic endothelium in tumor-draining lymph nodes from animals and patients, these results indicate that targeting integrin α4β1 or VCAM to inhibit the interactions of tumor cells with the lymph node microenvironment may be an effective strategy to suppress tumor metastasis.Tumor metastases are a leading cause of cancer-related mortality and morbidity, and both tumor cell intrinsic and extrinsic factors promote metastasis (1–4). Metastatic spread occurs primarily via lymphatic and hematogenous routes, and the presence of metastases in tumor draining lymph nodes is an accurate predictor of poor outcome in many types of tumors (5, 6). To further refine therapy for cancer patients, studies that define the mechanisms that promote tumor metastasis to lymph nodes could lead to novel therapeutic regimens that could improve clinical outcomes for cancer patients.In primary tumors, lymphangiogenesis, the growth of new lymphatic vessels, is strongly correlated with lymph node and distant metastasis. Increased expression of the lymphangiogenic factors VEGF-A, VEGF-C, or VEGF-D in tumors correlates closely with increased incidence of regional lymph node metastases in both humans and animals (7–9). Accordingly, systemic administration of antagonists of the VEGF-C receptor, VEGF-R3, blocked primary tumor lymphangiogenesis and metastasis (10–12).VEGF-C stimulates the expression of integrin α4β1, which promotes lymphatic endothelial cell (LEC) adhesion and invasion, leading to tumor-associated lymphangiogenesis (13). VEGF-C–mediated signaling stimulates LEC invasion and survival during lymphangiogenesis, as VEGF-R3 activates PI3K/v-akt murine thymoma viral oncogene homolog 1 (Akt) and mammalian target of rapamycin (mTOR) signaling pathways (14, 15). VEGF–VEGF-R3 signaling thus plays an important role in tumor lymphangiogenesis.Lymphangiogenesis occurs not only with primary tumors but also in tumor draining lymph nodes, where it is associated with increased tumor metastasis (16–18). However, it is unclear whether lymph node lymphangiogenesis plays an independent role in promoting tumor metastasis. Here we present the unique findings that integrin α4β1 is a biomarker of tumor-draining lymph nodes in animals and patients and that lymph node metastases depend on PI3Kα-mediated α4β1 activation in lymph node lymphatic endothelium. Once activated, α4β1 promotes lymph node lymphangiogenesis and facilitates adhesion of VCAM-1⁺ metastatic tumor cells within lymph nodes, thereby promoting tumor spread.