Gina M. Howell

RAS Mutations in Thyroid Cancer

In recent years, our understanding of the genetic alterations underlying thyroid oncogenesis has greatly expanded. The use of molecular markers, including RAS, in the management of thyroid carcinoma is also increasing. This review summarizes the current literature surrounding RAS and discusses its potential as a diagnostic and prognostic indicator in the management of thyroid cancer.

Augmenting Autophagy to Treat Acute Kidney Injury during Endotoxemia in Mice

Objective To determine that 1) an age-dependent loss of inducible autophagy underlies the failure to recover from AKI in older, adult animals during endotoxemia, and 2) pharmacologic induction of autophagy, even after established endotoxemia, is of therapeutic utility in facilitating renal recovery in aged mice. Design Murine model of endotoxemia and cecal ligation and puncture (CLP) induced acute kidney injury (AKI). Setting Academic research laboratory. Subjects C57Bl/6 mice of 8 (young) and 45 (adult) weeks of age. Intervention Lipopolysaccharide (1.5 mg/kg), Temsirolimus (5 mg/kg), AICAR (100 mg/kg). Measurements and Main Results: Herein we report that diminished autophagy underlies the failure to recover renal function in older adult mice utilizing a murine model of LPS-induced AKI. The administration of the mTOR inhibitor temsirolimus, even after established endotoxemia, induced autophagy and protected against the development of AKI. Conclusions These novel results demonstrate a role for autophagy in the context of LPS-induced AKI and support further investigation into like interventions that have potential to alter the natural history of disease.

Molecular mediators of metastasis in head and neck squamous cell carcinoma.

The presence of regional metastasis in patients with head and neck squamous cell carcinoma (HNSCC) is a common and adverse event associated with poor prognosis and high mortality. Although significant improvements in standard therapies have increased the efficacy of local tumor management, the high incidence of tumor recurrence has resulted in limited improvements in overall survival rates. Understanding the molecular mechanisms that mediate HNSCC invasion and metastasis may enable identification of novel therapeutic targets for the prevention and management of tumor dissemination.

Delay to Therapeutic Interventional Radiology Postinjury: Time Is of the Essence

OBJECTIVE Hemorrhage remains a leading cause of early death in injured patients, and definitive control of bleeding remains a fundamental principle of trauma management. Therapeutic interventional radiology (IR) procedures have increasingly become essential in the acute management of traumatic injury. The importance of time to control of hemorrhage for therapeutic IR procedures has not been adequately characterized. METHODS A retrospective analysis was performed by using data derived from the National Trauma Data Bank, version 7.1. Inclusion criteria included the following: adult, hypotensive patients, scene admission, patients who underwent early therapeutic IR vascular occlusive procedures within in 3 hours of admission at a level I or II designated trauma center (n = 1,748). Exclusion criteria included intracranial or venous occlusion procedures, patients who underwent any abdominal, thoracic, vascular, or intracranial operative procedures throughout their entire hospital stay. Logistic regression analysis was used to analyze the independent mortality risk associated with DELAY to IR procedures after controlling for important confounders. RESULTS The majority of patients who died did so within the first 48 hours from injury (80%). Regression analysis revealed that DELAY to IR was independently associated with more than a twofold higher risk of mortality (odds ratio 2.7, 95% confidence interval 1.6-4.9, p < 0.001). For every hour delay, the risk of mortality increased by 47%. These findings were independent of injury mechanism and most pertinent to level I trauma centers. CONCLUSION In hemodynamically unstable trauma patients undergoing therapeutic catheter-based IR procedures, delay to IR was independently associated with more than a twofold higher risk of mortality. These results suggest that therapeutic IR procedures should be performed as expeditiously as possible and held to the same dogma as applied to definitive operative control of hemorrhage.

Combined Inhibition of PLCγ-1 and c-Src Abrogates Epidermal Growth Factor Receptor–Mediated Head and Neck Squamous Cell Carcinoma Invasion

Purpose: Mortality from head and neck squamous cell carcinoma (HNSCC) is usually associated with locoregional invasion of the tumor into vital organs, including the airway. Understanding the signaling mechanisms that abrogate HNSCC invasion may reveal novel therapeutic targets for intervention. The purpose of this study was to investigate the efficacy of combined inhibition of c-Src and PLCγ-1 in the abrogation of HNSCC invasion. Experimental Design: PLCγ-1 and c-Src inhibition was achieved by a combination of small molecule inhibitors and dominant negative approaches. The effect of inhibition of PLCγ-1 and c-Src on invasion of HNSCC cells was assessed in an in vitro Matrigel-coated transwell invasion assay. In addition, the immunoprecipitation reactions and in silico database mining was used to examine the interactions between PLCγ-1 and c-Src. Results: Here, we show that inhibition of PLCγ-1 or c-Src with the PLC inhibitor U73122 or the Src family inhibitor AZD0530 or using dominant-negative constructs attenuated epidermal growth factor (EGF)–stimulated HNSCC invasion. Furthermore, EGF stimulation increased the association between PLCγ-1 and c-Src in HNSCC cells. Combined inhibition of PLCγ-1 and c-Src resulted in further attenuation of HNSCC cell invasion in vitro. Conclusions: These cumulative results suggest that PLCγ-1 and c-Src activation contribute to HNSCC invasion downstream of EGF receptor and that targeting these pathways may be a novel strategy to prevent tumor invasion in HNSCC.

CaMKIV-Dependent Preservation of mTOR Expression Is Required for Autophagy during Lipopolysaccharide-Induced Inflammation and Acute Kidney Injury

Autophagy, an evolutionarily conserved homeostasis process regulating biomass quantity and quality, plays a critical role in the host response to sepsis. Recent studies show its calcium dependence, but the calcium-sensitive regulatory cascades have not been defined. In this study, we describe a novel mechanism in which calcium/calmodulin-dependent protein kinase IV (CaMKIV), through inhibitory serine phosphorylation of GSK-3β and inhibition of FBXW7 recruitment, prevents ubiquitin proteosomal degradation of mammalian target of rapamycin (mTOR) and thereby augments autophagy in both the macrophage and the kidney. Under the conditions of sepsis studied, mTOR expression and activity were requisite for autophagy, a paradigm countering the current perspective that prototypically, mTOR inhibition induces autophagy. CaMKIV–mTOR-dependent autophagy was fundamentally important for IL-6 production in vitro and in vivo. Similar mechanisms were operant in the kidney during endotoxemia and served a cytoprotective role in mitigating acute kidney injury. Thus, CaMKIV–mTOR-dependent autophagy is conserved in both immune and nonimmune/parenchymal cells and is fundamental for the respective functional and adaptive responses to septic insult.

Calcium supplementation during sepsis exacerbates organ failure and mortality via calcium/calmodulin-dependent protein kinase kinase signaling.

Background:Calcium plays an essential role in nearly all cellular processes. As such, cellular and systemic calcium concentrations are tightly regulated. During sepsis, derangements in such tight regulation frequently occur, and treating hypocalcemia with parenteral calcium administration remains the current practice guideline. Objective:We investigated whether calcium administration worsens mortality and organ dysfunction using an experimental murine model of sepsis and explored the mechanistic role of the family of calcium/calmodulin-dependent protein kinases in mediating these physiological effects. To highlight the biological relevance of these observations, we conducted a translational study of the association between calcium administration, organ dysfunction, and mortality among a cohort of critically ill septic ICU patients. Design:Prospective, randomized controlled experimental murine study and observational clinical cohort analysis. Setting:University research laboratory and eight ICUs at a tertiary care center. Patients:A cohort of 870 septic ICU patients. Subjects:C57Bl/6 and CaMKK−/− mice. Interventions:Mice underwent cecal ligation and puncture polymicrobial sepsis and were administered with calcium chloride (0.25 or 0.25 mg/kg) or normal saline. Measurements and Main Results:Administering calcium chloride to septic C57Bl/6 mice heightened systemic inflammation and vascular leak, exacerbated hepatic and renal dysfunction, and increased mortality. These events were significantly attenuated in CaMKK−/− mice. In a risk-adjusted analysis of septic patients, calcium administration was associated with an increased risk of death, odds ratio 1.92 (95% CI, 1.00−3.68; p = 0.049), a significant increase in the risk of renal dysfunction, odds ratio 4.74 (95% CI, 2.48−9.08; p < 0.001), and a significant reduction in ventilator-free days, mean decrease 3.29 days (0.50−6.08 days; p = 0.02). Conclusions:Derangements in calcium homeostasis occur during sepsis that is sensitive to calcium administration. This altered calcium signaling, transduced by the calmodulin-dependent protein kinase kinase cascade, mediates heightened inflammation and vascular leak that culminates in elevated organ dysfunction and mortality. In the clinical management of septic patients, calcium supplementation provides no benefit and may impose harm.

Necrotizing Soft Tissue Infections

BACKGROUND Necrotizing soft tissue infections (NSTI) are an uncommon, but aggressive, problem with a potential for high morbidity and mortality rates. Establishing the diagnosis can be the major challenge. METHOD Review of pertinent English-language literature. RESULTS Early, aggressive surgical debridement combined with empiric broad-spectrum antibiotic therapy form the cornerstones of management. Novel therapeutic strategies such as hyperbaric oxygen, intravenous immunoglobulin, extracorporeal plasma treatment, and drotrecogin alfa (activated), also have been described, although their roles remain ill-defined. CONCLUSION Early diagnosis, despite its difficulties, is essential to guide the implementation of appropriate life-saving therapies. Understanding the microbiology of NSTI is important, not only to guide antibiotic therapy, but also to provide insight into the pathogenesis of the disease that will permit the future development of rationally targeted interventions.

CaMKIα Regulates AMP Kinase–Dependent, TORC-1–Independent Autophagy during Lipopolysaccharide-Induced Acute Lung Neutrophilic Inflammation

Autophagy is an evolutionarily conserved cytoplasmic process regulated by the energy rheostats mammalian target of rapamycin and AMP kinase (AMPK) that recycles damaged or unused proteins and organelles. It has been described as an important effector arm of immune cells. We have shown that the cytoplasmically oriented calcium/calmodulin-dependent protein kinase (CaMK)Iα regulates the inflammatory phenotype of the macrophage (Mϕ). In this study, we hypothesize that CaMKIα mediates Mϕ autophagy. LPS induced autophagy in RAW 264.7 cells and murine peritoneal Mϕ that was attenuated with biochemical CaMK inhibition or CaMKIα small interfering RNA (siRNA). Inhibition of CaMKIα reduced LPS-induced p-Thr172AMPK and target of rapamycin complex-1 activity, and expression of a constitutively active CaMKIα but not a kinase-deficient mutant induced p-Thr172AMPK and autophagy that was attenuated by the AMPK inhibitor compound C. Coimmunoprecipitation and in vitro kinase assays demonstrated that CaMKIα activates AMPK, thereby inducing ATG7, which also localizes to this CaMKIα/AMPK complex. During LPS-induced lung inflammation, C57BL/6 mice receiving CaMKIαsiRNA displayed reduced lung and bronchoalveolar immune cell autophagy that correlated with reduced neutrophil recruitment, myeloperoxidase activity, and air space cytokine concentration. Independently inhibiting autophagy, using siRNA targeting the PI3K VPS34, yielded similar reductions in lung autophagy and neutrophil recruitment. Thus, a novel CaMKIα/AMPK pathway is rapidly activated in Mϕ exposed to LPS and regulates an early autophagic response, independent of target of rapamycin complex-1 inhibition. These mechanisms appear to be operant in vivo in orchestrating LPS-induced lung neutrophil recruitment and inflammation.

Surgeon volume and adequacy of thyroidectomy for differentiated thyroid cancer

INTRODUCTION We aimed to determine influence of surgeon volume on (1) frequency of appropriate initial surgery for differentiated thyroid cancer (DTC) and (2) completeness of resection. METHODS We reviewed all initial thyroidectomies (Tx; lobectomy and total) performed in a health system during 2011; surgeons were grouped by number of Tx cases per year. For patients with histologic DTC ≥ 1 cm, surgeon volume was correlated with initial extent of the operation, and markers of complete resection including uptake on I(123) prescan, thyrotropin-stimulated thyroglobulin levels, and I(131) dose administered. RESULTS Of 1,249 patients who underwent Tx by 42 surgeons, 29% had DTC ≥ 1 cm without distant metastasis. At a threshold of ≥ 30 Tx per year, surgeons were more likely to perform initial total Tx for DTC ≥ 1 cm (P = .01), and initial resection was more complete as measured by all 3 quantitative markers. For patients with advanced stage disease, a threshold of ≥ 50 Tx per year was needed before observing improvements in I(123) uptake (P = .004). CONCLUSION Surgeons who perform ≥ 30 Tx a year are more likely to undertake the appropriate initial operation and have more complete initial resection for DTC patients. Surgeon volume is an essential consideration in optimizing outcomes for DTC patients, and even higher thresholds (≥ 50 Tx/year) may be necessary for patients with advanced disease.