Misoo C. Ellison

Possible role of human herpesvirus 8 in the lymphoproliferative disorders in common variable immunodeficiency

Patients who have common variable immunodeficiency (CVID) and granulomatous/lymphocytic interstitial lung disease (GLILD) are at high risk for early mortality and B cell lymphomas. Infection with human herpes virus type 8 (HHV8), a B cell lymphotrophic virus, is linked to lymphoproliferative disorders in people who have secondary immunodeficiencies. Therefore, we determined the prevalence of HHV8 infection in CVID patients with GLILD. Genomic DNA isolated from peripheral blood mononuclear cells was screened by nested- and real time-quantitative PCR (QRT-PCR) for the presence of HHV8 genome. It was positive in 6/9 CVID patients with GLILD (CVID-GLILD), 1/21 CVID patients without GLILD (CVID-control), and no patients receiving intravenous gamma globulin (n = 13) or normal blood donors (n = 20). Immunohistochemistry (IHC) demonstrated expression of the latency-associated nuclear antigen-1 (LANA-1) in the biopsies of the lung, liver, and bone marrow of four patients with CVID-GLILD. One CVID-GLILD patient developed a B cell lymphoma during the course of the study. QRT-PCR demonstrated high copy number of HHV8 genome and IHC showed diffuse staining for LANA-1 in the malignant lymph node. HHV8 infection may be an important factor in the pathogenesis of the interstitial lung disease and lymphoproliferative disorders in patients with CVID.

Elevated serum melatonin is associated with the nocturnal worsening of asthma

BACKGROUND Increased airway inflammation at night contributes to the nocturnal worsening of asthma. In vitro studies have shown exogenous melatonin to be pro-inflammatory in asthma, but it is unknown whether endogenous melatonin levels are a controller of airway inflammation in nocturnal asthma. OBJECTIVE Our aim was to determine 24-hour patterns of serum melatonin and their relationship to overnight decline in physiology in subjects with nocturnal asthma, non-nocturnal asthma, and in healthy controls. METHODS Observational study of pulmonary physiology and melatonin levels in patients with nocturnal asthma (n = 7), non-nocturnal asthma (n = 13), and healthy controls (n = 11). Subjects maintained a constant sleep-wake regimen for 7 days. On day 8, serum melatonin was measured every 2 hours by radioimmunoassay and analyzed by cosinor modeling. The correlation between serum melatonin levels and overnight change in spirometry was evaluated by Spearmans rank correlation analysis. RESULTS In subjects with nocturnal asthma, peak melatonin levels were significantly elevated compared with healthy controls (67.6 +/- 5.0 pg/mL versus 53.5 +/- 4.0 pg/mL, P =.03). Melatonin acrophase was delayed in nocturnal asthma (02:54 versus 01:58 in healthy controls, P =.003, and 02:15 in non-nocturnal asthma, P =.01). In subjects with nocturnal asthma, increasing melatonin levels were significantly and inversely correlated with overnight change in FEV(1) (r = -.79, P =.04), a relationship that was not observed in non-nocturnal asthma or healthy controls. CONCLUSIONS Nocturnal asthma is associated with elevation and phase delay of peak serum melatonin levels. Elevated melatonin levels might contribute to the pathogenesis of nocturnal asthma.

Cognition, MRS neurometabolites, and MRI volumetrics in non-neuropsychiatric systemic lupus erythematosus: preliminary data.

Objective:To correlate cognitive dysfunction with structural and neurometabolic brain findings in patients with non-neuropsychiatric systemic lupus erythematosus (non-NPSLE). Background:Over 25% of non-NPSLE patients have cognitive dysfunction, but the cerebral basis of this observation is not well understood. Method:Seven patients with non-NPSLE and seven control subjects were given a series of neuropsychological tests and neuroimaging with magnetic resonance imaging and magnetic resonance spectroscopy. Analyses of cognitive function and structural and neurometabolic measures of the brain were performed. Results:Compared with controls, the non-NPSLE patients were significantly impaired on a global cognitive impairment index (CII). No significant differences between the groups were found in choline/creatine (Ch/Cr), N-acetylaspartic acid/Cr, or hippocampal volumes. Ch/Cr was highly associated with CII across the sample. Conclusions:This is the first study to correlate cognitive impairment with an increase in Ch/Cr ratio among patients with SLE. These results, although preliminary, suggest that changes in cerebral white matter may be important in determining the subtle cognitive impairment that may occur in patients with SLE, even in the absence of neuropsychiatric symptoms.

Macrophages Kill Human Papillomavirus Type 16 E6-Expressing Tumor Cells by Tumor Necrosis Factor Alpha- and Nitric Oxide-Dependent Mechanisms

ABSTRACT The expression of adenovirus serotype 2 or 5 (Ad2/5) E1A sensitizes cells to killing by NK cells and activated macrophages, a property that correlates with the ability of E1A to bind the transcriptional coadaptor proteins p300-CBP. The E6 oncoproteins derived from the high-risk human papillomaviruses (HPV) interact with p300 and can complement mutant forms of E1A that cannot interact with p300 to induce cellular immortalization. Therefore, we determined if HPV type 16 (HPV16) E6 could sensitize cells to killing by macrophages and NK cells. HPV16 E6 expression sensitized human (H4 and C33A) and murine (MCA-102) cell lines to lysis by macrophages but not by NK cells. The lysis of cells that expressed E6 by macrophages was p53 independent but dependent on the production of tumor necrosis factor alpha (TNF-α) or nitric oxide (NO) by macrophages. Unlike cytolysis assays with macrophages, E6 expression did not significantly sensitize cells to lysis by the direct addition of NO or TNF-α. Like E1A, E6 has been reported to sensitize cells to lysis by TNF-α by inhibiting the TNF-α-induced activation of NF-κB. We found that E1A, but not E6, blocked the TNF-α-induced activation of NF-κB, an activity that correlated with E1A-p300 binding. In summary, Ad5 E1A and HPV16 E6 sensitized cells to lysis by macrophages. Unlike E1A, E6 did not block the ability of TNF-α to activate NF-κB or sensitize cells to lysis by NK cells, TNF-α, or NO. Thus, there appears to be a spectrum of common and unique biological activities that result as a consequence of the interaction of E6 or E1A with p300-CBP.

Surface Enhanced Laser Desorption/Ionization (SELDI) Time-of-Flight Mass Spectrometry to Identify Patients with Chronic Obstructive Pulmonary Disease

There are currently no blood tests to identify the majority of smokers at risk for chronic obstructive pulmonary disease (COPD). We used plasma protein profiles from surface enhanced laser desorption/ionization (SELDI) time-of-flight mass spectrometry to identify a panel of protein biomarkers that can distinguish patients with COPD from closely matched controls. Plasma was obtained from 30 COPD subjects and 30 controls matched for age, sex, and smoking history. Plasma protein profiles were generated using Cum2+-immobilized metal affinity capture (IMAC) and strong anion exchanger (Q10) protein chips. Classification and regression tree (CART) analysis identified a panel of 5 biomarkers using the IMAC protein chip that could distinguish COPD patients from controls with sensitivity and specificity of 91.67% and 88.33%, respectively. The 10-fold cross-validation yielded 81.67% sensitivity and 81.67% specificity. This demonstrates the feasibility of using SELDI as a diagnostic test for COPD; however, larger cohorts will be needed to validate these biomarkers and determine their predictive value longitudinally.

Feasibility of colocating dental hygienists into medical practices

OBJECTIVES To test the feasibility of colocating registered dental hygienists (RDHs) into medical practices and to evaluate parent/caregiver oral health characteristics. METHODS From December 2008 to April 2009, we colocated five RDHs into five medical practices identified for their service to low-income children. Dual-function exam rooms were built in each office. Caregiver-child dyads were recruited from the practices for program evaluation. We used both qualitative (key informant interviews) and quantitative (survey) methods to evaluate the project. Feasibility was measured by assessment of RDH and practice factors that facilitated and/or created barriers to colocation, sustainability of services 5 years after colocation, and caregiver satisfaction with services. Caregiver oral health knowledge, attitudes, beliefs, and behaviors were also measured. RESULTS Over 27 months, five part-time RDHs provided care to 2,071 children. Children of caregiver-child dyads (n = 583) recruited for evaluation were young (mean age = 1.8 years), white (46 percent), non-Hispanic (56 percent), and publicly insured (68 percent Medicaid/11 percent State Childrens Health Insurance Plan). Key informant interviews revealed various factors that facilitated and created barriers to program adoption, implementation, and sustainability. Most barriers were overcome. Five RDHs remained in the practices 2 years after program initiation and four remained after 5 years. At 1 year, 27 percent of caregiver-child dyads returned for evaluation and were highly satisfied with services. Caregivers reported favorable oral health characteristics and few barriers to receiving preventive dental care at baseline and 1-year follow-up. CONCLUSIONS Colocating RDHs into medical practices is feasible and an innovative model to provide preventive oral health services to disadvantaged children.

Impact of an interprofessional oral health education program on health care professional and practice behaviors: a RE-AIM analysis

Background Early childhood caries is the most common chronic childhood condition and largely preventable. Access to oral health preventive services (OHPS) for children at risk for caries is suboptimal and could be expanded if they were provided by non-dental professionals. Many state Medicaid programs in the USA now reimburse non-dental professionals for OHPS but require that they receive oral health education (OHE) to be reimbursed. Few OHE programs have been evaluated. Methods We evaluated the impact of Colorado’s OHE program on professional- and practice-level behaviors regarding the provision of OHPS to children by measuring its reach, effectiveness, adoption, implementation, and maintenance (ie, using the Reach Effectiveness Adoption Implementation Maintenance [RE-AIM] framework) with Medicaid claims data, online surveys, and key informant interviews. Results From 2009 to 2012, the proportion of young, low-income children receiving OHPS from a medical professional increased 16-fold. We surveyed 703 OHE participants; post-OHE response rates were 61% at 12 months, 34% at 24 months (2009 participants), and 39% at 12 months (2011 participants). Respondents reported confidence in providing OHPS; favorable oral health knowledge, attitudes, and beliefs; and were providing OHPS to most eligible children. Approximately half of the practices had initiated practice-level changes to support program implementation and maintenance. Few barriers were reported to care. Eighteen interviewees reported factors facilitating program diffusion, which included quality materials, community need, and reimbursement; barriers included lack of time to provide services, resources to purchase supplies, and referral dentists. Conclusion This evaluation of a state interprofessional OHE program shows evidence of program diffusion and identifies facilitating factors and barriers to having medical professionals provide OHPS.

RESPONDER CELL FREQUENCY ESTIMATION AND BINOMIAL THREE-LEVEL NONLINEAR MIXED EFFECTS MODEL IN LIMITING DILUTION ASSAYS

Abstract Responder cell frequencies (RCF), which describe vaccine-boosted immune responses in herpes zoster (HZ) prevention studies, have been estimated by using limiting dilution assays (LDA). The theoretical linearity assumption between the logarithm of the proportion of nonresponding wells (s) and the cell concentration (N) (or dilution level) in LDA, based on the single-hit Poisson model, is often violated with observed data resulting in biased estimates of RCF. In this article, the Poisson assumption is modified by applying a mixture of Poisson and gamma distributions, resulting in a negative binomial assumption, which presents a better fit between s and N. In LDA for HZ prevention studies, binary responses (responder or non-responder wells) are measured repeatedly at different cell concentrations and over time. To account for the correlation between responses to varying dilution levels from individuals, and the correlation between repeated assays of individuals over time simultaneously, a binomial three-level nonlinear mixed-effects model is proposed. For parameter estimation, a maximum likelihood method is applied via adaptive Gaussian quadrature. There is a lack of non-Gaussian multilevel nonlinear mixed-effects software, which can execute such a complicated fit. In this article, an algorithm for the three-level nonlinear mixed-effects model, which can be inserted into the code in the SAS® procedure NLMIXED, is suggested.

LUNG INFLATION WITH DIRECT INJECTION OF AGAROSE: A TECHNIQUE FOR SIMULTANEOUS MOLECULAR AND MORPHOMETRIC MEASUREMENTS

The usual methods for preparing lungs for morphologic study involve the instillation of fixatives that modify proteins and RNA such that the tissue is unsuitable for molecular studies. To develop a technique suitable for molecular studies, pieces of adult rat lungs were infiltrated with agarose, glutaraldehyde, or paraformaldehyde and the consistency of alveolar inflation was compared to lungs inflated with 10% formalin. Only direct injection with 1% agarose resulted in comparable inflation of lung tissue and preserved RNA and protein. Thus, this technique enables simultaneous molecular and morphometric analysis of the lung on small pieces of lung tissue in heterogeneous lung diseases.

79 Relationship between cognition and immune functions in chronic fatigue syndrome patients

This study examined associations between cognitive functioning and immune measures (cytokine, complement) in chronic fatigue syndrome (CFS) patients. Methods. Eighteen CFS patients (mean length diagnosis = 73.1 months, SEM = 12.0) and 8 healthy controls completed measures of cognition (i.e., attention, sequencing, verbal and nonverbal learning, memory, and verbal fluency), complement activity, and cytokine activity (CFS subjects only). Results. Using demographically corrected t scores, the CFS patients performed worse than controls on Trailmaking Test B, Digit Symbol and the Stroop Color Word Test. Complement activity measured by C5a was significantly higher in CFS participants compared to controls, however, no differences were found between the groups on C3a, C4a or ECP. Cytokine data were not collected in control subjects for comparison. Several significant correlations between cognitive performance and immune measures were found for CFS participants. Increased IL-1b was related to higher scores on BVMT learning (r = .86, p = .025) and decline on Letter Fluency (r = .84, p = .036). Higher levels of IL-6 were related to better performance on Trailmaking Test A (r = .56, p = .049) and Trailmaking Test B (r = .58, p = .034), and higher levels of IFN-a were related to better performance on BVMT Recall (r = .56, p = .046). In CFS patients only, none of the complement activity was associated with cognitive test scores. In control subjects only, increased C3a was associated with lower Animal Fluency (r = .72, p = .043) lower BVMT Delay (r = .077, p = .025) and improved Trailmaking Test A (r = .816, p = .013). Higher C5a was associated with better Animal Fluency (r = .71, p = .05). Higher ECP was associated with lower Letter Fluency (r = .756, p = .03) and better BVMT learning (r = .84, p = .008). Conclusions. In CFS participants, increased IL-1b was related to lower verbal fluency. In contrast, better performance in visuomotor speed, visuomotor sequencing, and visual learning were related to higher levels of IL-1b, IL6 and IFN-a. These results suggest both positive and negative relationships between immune and cognitive functions in CFS patients. There were no significant relationships within the CFS group between cognition and complement activity. However, in control subjects, associations between complement activity and verbal fluency and verbal and nonverbal learning/memory scores suggest continued evaluation in this area is warranted. Overall, continued analysis with a larger sample size of CFS and control subjects may be useful in better understanding the mechanisms underlying the relationships between immune and cognitive functions.