Global Health and Emergency Care: A Resuscitation Research Agenda—Part 2

At the 2013 Academic Emergency Medicine global health consensus conference, a breakout session to develop a research agenda for resuscitation was held. Two articles are the result of that discussion. This second article addresses data collection, management, and analysis and regionalization of postresuscitation care, resuscitation programs, and research examples around the world and proposes a strategy to strengthen resuscitation research globally. There is a need for reliable global statistics on resuscitation, international standardization of data, and development of an electronic standard for reporting data. Regionalization of postresuscitation care is a priority area for future research. Large resuscitation clinical research networks are feasible and can give valuable data for improvement of service and outcomes. Low‐cost models of population‐based research, and emphasis on interventional and implementation studies that assess the clinical effects of programs and interventions, are needed to determine the most cost‐effective strategies to improve outcomes. The global challenge is how to adapt research findings to a developing world situation to have an effect internationally.     

A high-efficiency Cre/loxP-based system for construction of adenoviral vectors

Adenovirus (Ad) vectors provide a highly efficient means of mammalian gene transfer and are widely used for high-level protein expression in mammalian cells, as recombinant vaccines and for gene therapy. A commonly used method for constructing Ad vectors relies on in vivo homologous recombination between two Ad DNA-containing bacterial plasmids cotransfected into 293 cells. While the utility of this two-plasmid approach is well established, its efficiency is low owing to the inefficiency of homologous recombination. To address this, we have developed an improved method for Ad vector construction based on Cre-mediated site-specific recombination between two bacterial plasmids, each bearing a loxP site. Ad vectors are generated as a result of Cre-mediated site-specific recombination between the two plasmids after their cotransfection into 293 cells expressing Cre recombinase. The frequency of Ad vector rescue by Cre-mediated site-specific recombination is significantly higher (approximately 30-fold) than by in vivo homologous recombination. The efficiency and reliability of this method should greatly simplify and expedite the construction of recombinant Ad vectors for mammalian gene transfer. Ad vectors are commonly constructed by homologous recombination between two plasmids cotransfected into 293 cells. This method has numerous advantages but results in low numbers of plaques owing to inefficient recombination. We have developed an improved method based on Cre-mediated site-specific recombination, which results in vector rescue at frequencies approximately 30-fold higher than by homologous recombination. This method should greatly simplify and expedite the construction of recombinant Ad vectors for mammalian gene transfer.     

An immune-sympathetic neuron communication axis guides adipose tissue browning in cancer-associated cachexia

Cancer-associated cachexia (CAC) is a hypermetabolic syndrome characterized by unintended weight loss due to the atrophy of adipose tissue and skeletal muscle. A phenotypic switch from white to beige adipocytes, a phenomenon called browning, accelerates CAC by increasing the dissipation of energy as heat. Addressing the mechanisms of white adipose tissue (WAT) browning in CAC, we now show that cachexigenic tumors activate type 2 immunity in cachectic WAT, generating a neuroprotective environment that increases peripheral sympathetic activity. Increased sympathetic activation, in turn, results in increased neuronal catecholamine synthesis and secretion, β-adrenergic activation of adipocytes, and induction of WAT browning. Two genetic mouse models validated this progression of events. 1) Interleukin-4 receptor deficiency impeded the alternative activation of macrophages, reduced sympathetic activity, and restrained WAT browning, and 2) reduced catecholamine synthesis in peripheral dopamine β-hydroxylase (DBH)–deficient mice prevented cancer-induced WAT browning and adipose atrophy. Targeting the intraadipose macrophage-sympathetic neuron cross-talk represents a promising therapeutic approach to ameliorate cachexia in cancer patients.

Cancer-associated cachexia (CAC) is an energy balance disorder causing unintended loss of body weight due to depletion of white adipose tissue (WAT) and skeletal muscle. This multiorgan and multifactorial syndrome affects up to 80% of cancer patients and is responsible for more than 20% of cancer-associated deaths (1). CAC impedes the effectiveness of anticancer therapies and drastically lowers patients’ quality of life (2).A long list of tumor-borne, often proinflammatory factors, including interleukin-6 (IL-6) (3), parathyroid hormone–related protein (PTHrP) (4), leukemia inhibitory factor (LIF) (5), zinc α-glycoprotein (6), or growth differentiation factor-15 (GDF-15) (7), trigger CAC in mouse models. However, the signaling cascades and catabolic mechanisms that lead to adipose- and muscle tissue wasting remain insufficiently understood (8, 9). IL-6 and PTHrP are among the best studied of these “cachexokines.” Their presence or absence is decisive for the development of CAC in cancer patients and animal models (4, 10–13). Thus, treatment with neutralizing antibodies against IL-6, the IL-6 receptor, or PTHrP ameliorates CAC in various mouse models of CAC (3, 4, 14, 15).CAC-associated WAT atrophy results from a metabolic switch toward decreased lipid synthesis and excessive degradation of lipid stores via enhanced triglyceride degradation (lipolysis) (9, 16). Induced lipolysis is observed in both humans and mice with CAC (17, 18). The absence of metabolic lipases at least partially ameliorates cachexia in murine cancer models (19). The metabolic or catabolic fates of lipolytic products, namely fatty acids (FAs) and glycerol, have not been fully clarified. These may provide energy and/or biosynthetic substrates for cancer cells to promote tumor growth or can be reesterified in WAT, creating an adenosine-triphosphate (ATP)-consuming futile metabolic cycle. Both of these pathways would contribute to the eventual loss of WAT during CAC (20).Another important catabolic pathway in CAC involves the direct oxidation of FAs and glycerol in adipose tissue. This process is promoted by the conversion of white to beige adipocytes called “WAT browning.” During WAT browning, adipocytes adopt a multilocular lipid droplet morphology; express genes that are typical for brown adipocytes, such as uncoupling protein-1 (UCP-1); exhibit elevated substrate oxidation rates; and dissipate energy as heat (21). WAT browning occurs in carcinogen-induced cancer models and genetically engineered mouse models as well as syngeneic and xenogeneic transplant models of murine cancers (3, 4, 22) and depends on the presence of cachexokines. WAT browning also occurs in humans suffering CAC or severe burn trauma (3, 23–25), but the cellular and molecular mechanisms underlying catabolic WAT remodeling in CAC remain unclear.Here, we demonstrate that a macrophage-sympathetic neuron signaling axis generates a high β-adrenergic tone resulting in beige adipogenesis, increased lipid degradation, and WAT atrophy in murine models of CAC. This mechanism triggering hypermetabolism in CAC may offer targets for prevention or treatment of the disease.     

Actions of recombinant human γ-interferon and tumor necrosis factor α on the proliferation and osteoblastic characteristics of human trabecular bone cells in vitro

Using cultured human osteoblast-like cells, we studied the effects of tumor necrosis factor (TNF) and recombinant human γ-interferon (γ-IFN) on osteoblast growth and function, and demonstrated that TNF stimulated bone cell proliferation and prostaglandin production while inhibiting 1,25-(OH)₂D₃—stimulated alkaline phosphatase activity and osteocalcin release. In contrast, γ-IFN inhibited proliferation and stimulated alkaline phosphatase activity of the cells, while inhibiting 1,25-(OH)₂D₃—stimulated osteocalcin production and having variable effects on the release of prostaglandins, depending on the presence of other factors. Our results suggest that TNF and γ-IFN can act directly on bone-forming cells to affect both their proliferation and their differentiated function, and that changes in the ability of cells to produce these factors in disease states may contribute to alterations in the integrity of connective tissue matrices.     

A non-invasive, relocatable stereotactic frame for fractionated radiotherapy and multiple imaging.

A non-invasive relocatable stereotactic frame has been developed for fractionated stereotactic external beam radiotherapy (SRT) by linear accelerator. Fixation is via the upper dentition and tests of repositioning have shown mean antero-posterior and lateral displacements of 1.0 mm. The relocatable frame allows accurate 3-dimensional target localisation by CT, MRI, PET and cerebral angiography and precise isocentric positioning for radiotherapy. This method of immobilisation is ideal for fractionated SRT as well as for other techniques requiring high precision localisation and treatment delivery.     

Multimodal Percutaneous Intervention for Critical Venous Occlusive Disease

Critical deep venous thrombosis and occlusion constitutes a small percentage of patients with venous disease, who exhibit severe symptomatology. This study examined the results of multimodal percutaneous therapy for the treatment of complex critical venous thrombotic and occlusive disease. Twenty-five patients presented with critical venous thromboses or occlusions (11 with debilitating unilateral lower extremity edema causing ambulatory impairment, 2 with debilitating bilateral lower extremity edema, 3 with phlegmasia cerulea dolens, 2 with venous claudication, 2 with superior vena cava (SVS) syndrome with respiratory compromise, 4 with debilitating upper extremity edema, and 1 with renal insufficiency). Therapeutic modalities including thrombolysis, mechanical thrombectomy, percutaneous venoplasty and stent placement, temporary inferior vena cava filtration, and ultrasound guidance were used in all cases in conjunction with long-term systemic anticoagulation. The venous access site was determined by the anatomic location of the lesion and included popliteal, femoral, brachial, and lesser saphenous. Patients were followed with clinical exam and duplex surveillance. Resolution of symptoms was achieved in 18 of 25 patients (72%) and partial resolution occurred in 4 of 25 (16%). Failure of treatment identified as both lack of clinical response and evidence of continued venous thrombosis occurred 3 of 25 patients (12%). Restoration of arterial pulses and limb salvage was achieved in the three patients with phlegmasia cerulea dolens and acute limb-threatening ischemia. Both patients with SVC syndrome experienced resolution of respiratory compromise and facial edema. The mean length of follow-up was 11 ± 2.7 months. Complications included transfusion requirement (2), hematuria (2), retroperitoneal hematoma (1), and cellulitis (1). Acute critical venous thrombotic and occlusive disease is responsive to multimodal percutaneous treatment. The relief of pain and resolution of acutely life and limb-threatening conditions in this most severely symptomatic subset of patients represents the immediate goal of treatment.Presented at the Twenty-ninth Annual Meeting of the Peripheral Vascular Surgery Society, Anaheim, CA, June 4-5, 2004.