Proportion of CD4 and CD8 tumor infiltrating lymphocytes predicts survival in persistent/recurrent laryngeal squamous cell carcinoma.

Tumor infiltrating lymphocytes (TILs) have been shown to be an important prognostic factor in patients with previously untreated head and neck cancer. After organ preservation therapy for laryngeal cancer and subsequent persistence/recurrence, the prognostic value of TILs is unknown. Our goal was to determine if TILs have value as a prognostic biomarker in patients with surgically salvageable persistent/recurrent laryngeal squamous cell carcinoma. Levels of TILs were quantified on tissue microarrays from 183 patients undergoing salvage total laryngectomy for persistent/recurrent laryngeal cancer after radiation or chemoradiation between 1997 and 2014. Demographic and clinical data were abstracted. Immunohistology evaluation included CD4, CD8, PDL-1, p16, CD31, Vimentin, EGFR, and p53. Elevated levels of either CD8 or CD4 positive TILs were associated with improved disease specific survival (CD8: HR 0.46, 95% CI 0.24-0.88, CD4: HR 0.43; 95% CI 0.21-0.89) and disease free survival (CD8: HR 0.53, 95% CI 0.29-0.94, CD4: HR 0.52; 95% CI 0.27-0.99). Levels of CD8 (HR 0.74; 95% CI 0.47-1.17) or CD4 (HR 0.66; 95% CI 0.40-1.08) TILs were not significantly associated with overall survival. In bivariate analysis, patients with elevated CD4 and/or CD8 TILs had significantly improved disease specific survival (HR 0.42; 95% CI 0.21-0.83) and disease free survival (HR 0.45; 95% CI 0.24-0.84) compared to patients with low levels of CD4 and CD8. PDL-1, p16, CD31, Vimentin, EGFR, and p53 were not significant prognostic factors. On multivariate analysis, elevated CD8 TILs were associated with improved disease specific survival (HR 0.35; 95% CI 0.14-0.88, p = .02) and disease free survival (HR 0.41; 95% CI 0.17-0.96, p = .04). CD8, and possibly CD4, positive TILs are associated with favorable disease free and disease specific survival for recurrent/persistent laryngeal cancer.

In-hospital statin underutilization among high-risk patients: delayed uptake of the 2013 cholesterol guidelines in a U.S. cohort.

OBJECTIVES: Clinician utilization of the 2013 cholesterol lowering guidelines remains variable and unknown. We sought to examine statin prescribing patterns and compare rates among specialists who treat high-risk cardiovascular patients admitted to the hospital. METHODS: We retrospectively (via chart review) examined four specialty groups: (i) Cardiology, (ii) Cardiovascular or Vascular (CV) Surgery, (iii) Neurology, and (iv) Internal Medicine. Adult patients were included based on a discharge diagnosis of acute coronary syndrome, coronary artery bypass graft surgery, carotid endarterectomy, acute ischemic stroke, transient ischemic attack, or high-risk chest pain. Prescribing patterns were evaluated 6 months and 18 months after the release of the 2013 guidelines. High-intensity statin was defined as atorvastatin 40-80 mg or rosuvastatin 20-40 mg per day. RESULTS: 632 patients were included in our study. The following percentages of patients were discharged on high-intensity statin (6 months; 18 months): (i) Cardiology (80%; 85%), (ii) CV Surgery (52%, 65%), (iii) Neurology (59%; 66%), and (iv) Internal Medicine (45%; 48%). Among the four groups, Cardiology was the most likely to discharge patients on high-intensity statin (p < 0.001) in 2014 and in 2015. Cardiology, CV Surgery, and Neurology significantly increased the percentage of patients on high-intensity statin from pre-admission to time of discharge in both years. CONCLUSION: High-intensity statin therapy is underutilized among high-risk cardiovascular patients admitted to the hospital. Variations exist in prescribing patterns of different specialties who manage high-risk populations. This data can be used to test quality improvement interventions to improve rates of high-intensity statin utilization among high-risk patients prior to hospital discharge.

Effects of low dose metformin in adolescents with type I diabetes mellitus: a randomized, double-blinded placebo-controlled study.

BACKGROUND: Insulin resistance increases during adolescence in those with type 1 diabetes mellitus (T1DM), complicating glycemic control and potentially increasing cardiovascular disease (CVD) risk. Metformin, typically used in type 2 diabetes mellitus (T2DM), is a possible adjunct therapy in T1DM to help improve glycemic control and insulin sensitivity. OBJECTIVE: We hypothesized that metformin would improve metabolic parameters in adolescents with T1DM. DESIGN, SETTING, AND PARTICIPANTS: This randomized, double-blinded, placebo-controlled trial included 74 pubertal adolescents (ages: 13-20 yr) with T1DM. Participants were randomized to receive either metformin or placebo for 6 months. Glycated hemoglobin (HbA1c), insulin dose, waist circumference, body mass index (BMI), and blood pressure were measured at baseline, 3 and 6 months, with fasting lipids measured at baseline and 6 months. RESULTS: Total daily insulin dose, BMI z-score and waist circumference significantly decreased at 3 and 6 months compared to baseline within the metformin group, even among normal-weight participants. In the placebo group, total insulin dose and systolic blood pressure increased significantly at 3 months and total insulin dose increased significantly at 6 months. No significant change was observed in HbA1c at any time point between metformin and placebo groups or within either group. CONCLUSIONS: Low-dose metformin likely improves BMI as well as insulin sensitivity in T1DM adolescents, as indicated by a decrease in total daily insulin dose. The decrease in waist circumference indicates that fat distribution is also likely impacted by metformin in T1DM. Further studies with higher metformin doses and more detailed measurements are needed to confirm these results, their underlying mechanisms, and potential impact on CVD in T1DM youth.

The StatStrip glucose monitor is suitable for use during hyperinsulinemic euglycemic clamps in a pediatric population.

BACKGROUND: The hyperinsulinemic euglycemic clamp is the gold standard for assessment of insulin resistance and requires frequent, accurate measurements of blood glucose concentrations, typically utilizing the YSI 2300 STAT Plus™ glucose analyzer (YSI, Inc., Yellow Springs, OH). Despite its accuracy, the YSI has several limitations, including its cost, lengthy run time, need for trained personnel, frequent maintenance, and large blood volumes. Simpler hospital-grade hand-held glucose meters are now available but have not been validated for use in pediatric clamp settings. Our objective was to evaluate the accuracy, precision, and reliability of the StatStrip(®) (SS) hospital glucose monitoring system (Nova Biomedical, Waltham, MA) relative to the YSI 2300 STAT glucose analyzer in pediatric hyperinsulinemic euglycemic clamps. SUBJECTS AND METHODS: Four hundred sixty blood specimens drawn from 11 pediatric patients undergoing hyperinsulinemic euglycemic clamps were simultaneously analyzed by SS and YSI. Outcome measures included SS bias relative to YSI and glucose measurement precision on SS and YSI. RESULTS: The SS showed a slight positive bias of 0.75 ± 2.83 mg/dL versus the YSI. Percentage coefficients of variance for SS and YSI were 9.53% and 9.25%, respectively. Using a Bland-Altman plot, the limits of agreement were ± 5.7 mg/dL. The coefficient of repeatability for SS was 6.63; the coefficient of individual agreement between the YSI and SS was 0.995. CONCLUSIONS: The SS is a suitable replacement for the YSI in pediatric hyperinsulinemic euglycemic clamp studies, is easier to use, more cost-effective, and faster, and requires less blood. Future euglycemic clamp studies can consider utilizing this methodology.

Dysfunctional resident lung mesenchymal stem cells contribute to pulmonary microvascular remodeling.

Pulmonary vascular remodeling and oxidative stress are common to many adult lung diseases. However, little is known about the relevance of lung mesenchymal stem cells (MSCs) in these processes. We tested the hypothesis that dysfunctional lung MSCs directly participate in remodeling of the microcirculation. We employed a genetic model to deplete extracellular superoxide dismutase (EC-SOD) in lung MSCs coupled with lineage tracing analysis. We crossed (floxp)sod3 and mT/mG reporter mice to a strain expressing Cre recombinase under the control of the ABCG2 promoter. We demonstrated In vivo that depletion of EC-SOD in lung MSCs resulted in their contribution to microvascular remodeling in the smooth muscle actin positive layer. We further characterized lung MSCs to be multipotent vascular precursors, capable of myofibroblast, endothelial and pericyte differentiation in vitro. EC-SOD deficiency in cultured lung MSCs accelerated proliferation and apoptosis, restricted colony-forming ability, multilineage differentiation potential and promoted the transition to a contractile phenotype. Further studies correlated cell dysfunction to alterations in canonical Wnt/ß-catenin signaling, which were more evident under conditions of oxidative stress. Our data establish that lung MSCs are a multipotent vascular precursor population, a population which has the capacity to participate in vascular remodeling and their function is likely regulated in part by the Wnt/ß-catenin signaling pathway. These studies highlight an important role for microenviromental regulation of multipotent MSC function as well as their potential to contribute to tissue remodeling.

Isolation & characterization of Hoechst(low) CD45(negative) mouse lung mesenchymal stem cells.

Tissue resident mesenchymal stem cells (MSC) are important regulators of tissue repair or regeneration, fibrosis, inflammation, angiogenesis and tumor formation. Taken together these studies suggest that resident lung MSC play a role during pulmonary tissue homeostasis, injury and repair during diseases such as pulmonary fibrosis (PF) and arterial hypertension (PAH). Here we describe a technology to define a population of resident lung MSC. The definition of this population in vivo pulmonary tissue using a define set of markers facilitates the repeated isolation of a well-characterized stem cell population by flow cytometry and the study of a specific cell type and function.

Osteoblasts derived from induced pluripotent stem cells form calcified structures in scaffolds both in vitro and in vivo.

Reprogramming somatic cells into an ESC-like state, or induced pluripotent stem (iPS) cells, has emerged as a promising new venue for customized cell therapies. In this study, we performed directed differentiation to assess the ability of murine iPS cells to differentiate into bone, cartilage, and fat in vitro and to maintain an osteoblast phenotype on a scaffold in vitro and in vivo. Embryoid bodies derived from murine iPS cells were cultured in differentiation medium for 8­12 weeks. Differentiation was assessed by lineage-specific morphology, gene expression, histological stain, and immunostaining to detect matrix deposition. After 12 weeks of expansion, iPS-derived osteoblasts were seeded in a gelfoam matrix followed by subcutaneous implantation in syngenic imprinting control region (ICR) mice. Implants were harvested at 12 weeks, histological analyses of cell and mineral and matrix content were performed. Differentiation of iPS cells into mesenchymal lineages of bone, cartilage, and fat was confirmed by morphology and expression of lineage-specific genes. Isolated implants of iPS cell-derived osteoblasts expressed matrices characteristic of bone, including osteocalcin and bone sialoprotein. Implants were also stained with alizarin red and von Kossa, demonstrating mineralization and persistence of an osteoblast phenotype. Recruitment of vasculature and microvascularization of the implant was also detected. Taken together, these data demonstrate functional osteoblast differentiation from iPS cells both in vitro and in vivo and reveal a source of cells, which merit evaluation for their potential uses in orthopedic medicine and understanding of molecular mechanisms of orthopedic disease.

The pathology of bleomycin-induced fibrosis is associated with loss of resident lung mesenchymal stem cells that regulate effector T-cell proliferation.

Tissue-resident mesenchymal stem cells (MSCs) are important regulators of tissue repair or regeneration, fibrosis, inflammation, angiogenesis, and tumor formation. Here, we define a population of resident lung MSCs (luMSCs) that function to regulate the severity of bleomycin injury via modulation of the T-cell response. Bleomycin-induced loss of these endogenous luMSCs and elicited fibrosis (pulmonary fibrosis), inflammation, and pulmonary arterial hypertension (PAH). Replacement of resident stem cells by administration of isolated luMSCs attenuated the bleomycin-associated pathology and mitigated the development of PAH. In addition, luMSC modulated a decrease in numbers of lymphocytes and granulocytes in bronchoalveolar fluid and demonstrated an inhibition of effector T-cell proliferation in vitro. Global gene expression analysis indicated that the luMSCs are a unique stromal population differing from lung fibroblasts in terms of proinflammatory mediators and profibrotic pathways. Our results demonstrate that luMSCs function to protect lung integrity after injury; however, when endogenous MSCs are lost, this function is compromised illustrating the importance of this novel population during lung injury. The definition of this population in vivo in both murine and human pulmonary tissue facilitates the development of a therapeutic strategy directed at the rescue of endogenous cells to facilitate lung repair during injury.