Brain metastatic breast cancer (BMBC) is uniformly fatal and increasing in frequency. Despite its devastating outcome, mechanisms causing BMBC remain largely unknown. The mechanisms that implicate circulating tumor cells (CTCs) in metastatic disease, notably in BMBC, remain elusive. Here we characterize CTCs isolated from peripheral blood mononuclear cells (PBMCs) of patients with breast cancer, and also develop CTC lines from three of these patients. In epithelial cell adhesion molecule (EpCAM)–negative CTCs, we identified a potential signature of brain metastasis comprising “brain metastasis selected markers (BMSM)” HER2+/EGFR+/HPSE+/Notch1+. These CTCs—which are not captured by the CellSearch platform because of their EpCAM negativity—were analyzed for cell invasiveness and metastatic competency in vivo. CTC lines expressing the BMSM signature were highly invasive and capable of generating brain and lung metastases when xenografted in nude mice. Notably, increased brain metastatic capabilities, frequency, and quantitation were detected in EpCAM− CTCs overexpressing the BMSM signature. The presence of proteins of the BMSM CTC signature was also detected in the metastatic lesions of animals. Collectively, we provide evidence of isolation, characterization, and long-term culture of human breast cancer CTCs, leading to the description of a BMSM protein signature that is suggestive of CTC metastatic competency to the brain.

Importance There is limited information about the effect of erythropoietin or a high transfusion threshold in traumatic brain injury (TBI). Objective To compare the effects of erythropoietin and two transfusion thresholds (7 and 10 g/dl) on neurological recovery after TBI. Design Randomized trial using a factorial design to test: i.) whether erythropoietin would fail to improve favorable outcomes by 20%, and ii.) whether a transfusion threshold of >10 g/dl would increase favorable outcomes without increasing complications. Setting Neurosurgical intensive care units of two Houston level 1 trauma centers Participants Between May 2006 and August 2012, 200 patients with closed head injury who were unable to follow commands were enrolled within 6 hours of injury; 102 patients received erythropoetin and 98 received placebo. Erythropoetin or placebo was initially dosed daily for 3 days and then weekly for 2 more weeks (n=74) and then the 24h and 48h doses were dropped for the remainder (n=126). Ninety-nine and 101 patients were assigned to the 7g/dl and 10g/dl transfusion thresholds. Intervention Intravenous erythropoietin 500 IU/kg or saline per dose. Transfusion threshold maintained with packed red blood cell transfusion. Main Outcome Glasgow Outcome Scale dichotomized as favorable (good recovery and moderate disability) and unfavorable (severe disability, vegetative, or dead) at 6 months post-injury. Results There was no erythropoeitin-transfusion threshold interaction. Compared to placebo (favorable outcome rate: 34/89 [38.2%]; 95%CI=28.2-49.1%), both erythropoetin groups were futile (first dosing regimen: 17/35 [48.6%]; 95%CI=31.4-66.0%, p=0.13, and second dosing regimen: 17/57 [29.8%]; 95%CI=18.4-43.4%, p<0.001). Favorable outcome rates were 37/87 (42.5%) and 31/94 (33.0%) in the 7 and 10 g/dl threshold groups (95%CI for the difference = − 0.05 to 0.25, p=0.28). There was a higher incidence of thromboembolic events in the 10 g/dl threshold group (22/101 [21.8%] vs. 8/99 [8.1%], p=0.009). Conclusions and Relevance In patients with closed head injury, neither the administration of erythropoietin nor maintaining hemoglobin concentration > 10 g/dl resulted in improved neurological outcome at 6 months and the 10 g/dl threshold was associated with a higher incidence of adverse events.. These findings do not support either approach in this setting.

Microdialysis enables the chemistry of the extracellular interstitial space to be monitored. Use of this technique in patients with acute brain injury has increased our understanding of the pathophysiology of several acute neurological disorders. In 2004, a consensus document on the clinical application of cerebral microdialysis was published. Since then, there have been significant advances in the clinical use of microdialysis in neurocritical care. The objective of this review is to report on the International Microdialysis Forum held in Cambridge, UK, in April 2014 and to produce a revised and updated consensus statement about its clinical use including technique, data interpretation, relationship with outcome, role in guiding therapy in neurocritical care and research applications.Electronic supplementary materialThe online version of this article (doi:10.1007/s00134-015-3930-y) contains supplementary material, which is available to authorized users.

Dendritic cells are potent antigen-presenting cells that initiate and amplify immune responses. To determine whether dendritic cells participate in inflammatory reactions in amyotrophic lateral sclerosis (ALS), we examined mRNA expression of dendritic cell surface markers in individual sporadic ALS (sALS), familial ALS (fALS), and nonneurological disease control (NNDC) spinal cord tissues using semiquantitative and real-time reverse transcription polymerase chain reaction (RT-PCR). Immature (DEC205, CD1a) and activated/mature (CD83, CD40) dendritic cell transcripts were significantly elevated in ALS tissues. The presence of immature and activated/mature dendritic cells (CD1a(+) and CD83(+)) was confirmed immunohistochemically in ALS ventral horn and corticospinal tracts. Monocytic/macrophage/microglial transcripts (CD14, CD18, SR-A, CD68) were increased in ALS spinal cord, and activated CD68(+) cells were demonstrated in close proximity to motor neurons. mRNA expressions of the chemokine MCP-1, which attracts monocytes and myeloid dendritic cells, and of the cytokine macrophage-colony stimulating factor (M-CSF) were increased in ALS tissues. The MCP-1 protein was expressed in glia in ALS but not in control tissues and was increased in the CSF of ALS patients. Those patients who progressed most rapidly expressed significantly more dendritic transcripts than patients who progressed more slowly. These results support the involvement of immune/inflammatory responses in amplifying motor neuron degeneration in ALS.

ABSTRACTy-Glutamyl transpeptidase (GGT) is an ectoenzyme that catalyzes the first step in the cleavage of glutathione (GSH) and plays an essential role in the metabolism of GSH and GSH conjugates of carcinogens, toxins, and eicosanoids. To learn more about the role of GGT in metabolism in vivo, we used embryonic stem cell technology to generate GGT-deficient (GGTml/GGTml) mice. GGTdeficient mice appear normal at birth but grow slowly and by 6 weeks are about half the weight of wild-type mice. They are sexually immature, develop cataracts, and have coats with a gray cast. Most die between 10 and 18 weeks. Plasma and urine GSH levels in the GGTml/GGTml mice are elevated 6-fold and 2500-fold, respectively, compared with wild-type mice. Tissue GSH levels are markedly reduced in eye, liver, and pancreas.Plasma cyst(e)ine levels in GGTm'/GGTml mice are reduced to '20% of wild-type mice. Oral administration of Nacetylcysteine to GGTml/GGTml mice results in normal growth rates and partially restores the normal agouti coat color. These findings demonstrate the importance of GGT and the y-glutamyl cycle in cysteine and GSH homeostasis.