Dysregulation of neutrophil extracellular traps (NETs)-related genes in the pathogenesis of diabetic kidney disease - Results from bioinformatics analysis and translational studies.

The role of Neutrophil extracellular traps (NETs) in the immunopathogenesis of Diabetic Kidney Disease (DKD) remains elusive. We used a machine learning approach to identify differentially expressed genes (DEGs) associated with NETs in human DKD kidney biopsy datasets and validated the results using single-nucleus RNA sequencing datasets. The expressions of these candidate genes and related cytokines were verified in blood obtained from DKD patients. Three NETs-associated genes (ITGAM, ITGB2 and TLR7) were identified, which all showed significant upregulation in both glomerular and tubulointerstitial compartments in human DKD kidneys. DKD patients showed significantly higher number of activated neutrophils with increased ITGAM and ITGB2 expression, higher serum IL-6 but lower IL-10, compared to healthy controls (p all <0.01). This study suggests that dysregulation of NETs-associated genes ITGAM and ITGB2 are related to the pathogenesis of DKD, and may serve as novel diagnostic markers and therapeutic targets in DKD.

Early Changes in Tumor-Naive Cell-Free Methylomes and Fragmentomes Predict Outcomes in Pembrolizumab-Treated Solid Tumors.

Early kinetics of circulating tumor DNA (ctDNA) in plasma predict response to pembrolizumab but typically requires sequencing of matched tumor tissue or fixed gene panels. We analyzed genome-wide methylation and fragment-length profiles using cell-free methylated DNA immunoprecipitation and sequencing (cfMeDIP-seq) in 204 plasma samples from 87 patients before and during treatment with pembrolizumab from a pan-cancer phase II investigator-initiated trial (INSPIRE). We trained a pan-cancer methylation signature using independent methylation array data from The Cancer Genome Atlas to quantify cancer-specific methylation (CSM) and fragment-length score (FLS) for each sample. CSM and FLS are strongly correlated with tumor-informed ctDNA levels. Early kinetics of CSM predict overall survival and progression-free survival, independently of tumor type, PD-L1, and tumor mutation burden. Early kinetics of FLS are associated with overall survival independently of CSM. Our tumor-naïve mutation-agnostic ctDNA approach integrating methylomics and fragmentomics could predict outcomes in patients treated with pembrolizumab. SIGNIFICANCE: Analysis of methylation and fragment length in plasma using cfMeDIP-seq provides a tumor-naive approach to measure ctDNA with results comparable with a tumor-informed bespoke ctDNA. Early kinetics within the first weeks of treatment in methylation and fragment quantity can predict outcomes with pembrolizumab in patients with various advanced solid tumors. This article is featured in Selected Articles from This Issue, p. 897.

Effect of (poly)phenol-rich 'Daux Belan' apple supplementation on diet-induced obesity and glucose intolerance in C57BL/6NCrl mice.

Obesity is a state of metabolic dysfunction that can lead to dyslipidemia and impaired glucose homeostasis. Apple polyphenols have been shown to ameliorate dyslipidemia/metabolic dysfunction in humans. The influence of apple (poly)phenols on energy metabolism in high-fat (HF) diet-induced obese mice remains controversial. This study examined the effect of dietary supplementation of (poly)phenol-rich 'Daux Belan' apple (DB; 6.2 mg gallic acid equivalence (GAE)/mouse/day; 0.15% (poly)phenol) in the form of freeze-dried powder on glucose and lipid metabolism in male HF-fed C57BL/6NCrl mice, in comparison to low-(poly)phenol-containing 'Zestar' apple (Z; 0.4 mg GAE/mouse/day). Obesity, glucose intolerance, hypertriglyceridemia, and hepatic lipid vacuolation were induced by HF feeding while circulating cholesterol levels remained unchanged. DB apple supplementation did not protect against HF-induced body weight gain, hyperglycemia, hepatic triglyceride level elevation, and hepatic lipid vacuolation at the tested dosage. Future studies should be conducted with increased DB dosage and employ apple (poly)phenols supplemented in the form of extracts or sugar-free powder.

Double Trouble: Whole-Genome Doubling Distinguishes Early from Late Ovarian Cancer.

Dramatic differences in outcome between early- and late-stage high-grade serous ovarian cancer (HGSC) suggest perhaps distinct genetic origins due to differences in exposures to mutational processes. Evidence to support this hypothesis was recently reported by comparative analysis of copy-number signatures between early- and late-stage HGSCs. See related article by Cheng et al., p. 2911.

Cost-effectiveness of olaparib versus routine surveillance in the maintenance setting for patients with BRCA-mutated advanced ovarian cancer after response to first-line platinum-based chemotherapy in Singapore.

OBJECTIVE: To evaluate the cost-effectiveness of olaparib as a maintenance treatment versus routine surveillance (RS) in patients with BRCA mutated (BRCAm) advanced ovarian cancer (OC) following response to first-line platinum-based chemotherapy in Singapore. METHODS: A 4-health state partitioned survival model was developed to simulate the lifetime (50 years) incremental cost-effectiveness ratio (ICER) of olaparib versus RS from a healthcare payer perspective. Progression-free survival, time to second disease progression, and overall survival were estimated using SOLO-1 data and extrapolated beyond the trial period using parametric survival models. Any patient who remained progression-free at year 7 was assumed to be no longer at risk of progression. Mortality rates were based on all-cause mortality, adjusted based on BRCA1/2 mutation. Health state utilities and adverse event frequencies were from SOLO-1. Drug costs were from local public healthcare institutions. Healthcare resource usage and costs were from local clinician input and publications. A 3% discount rate was applied to costs and outcomes. Deterministic and probabilistic sensitivity analyses (PSA) were performed to assess the robustness of results. RESULTS: The base-case analysis of olaparib maintenance therapy versus RS resulted in an ICER of Singapore dollar (SGD) 19,822 per quality-adjusted life-year (QALY) gained. The ICER was most sensitive to variations in the discount rate. PSA demonstrated that olaparib had an 87% probability of being cost-effective versus RS at a willingness-to-pay of SGD 60,000 per QALY gained. CONCLUSION: Olaparib has a high potential of being a cost-effective maintenance treatment versus RS for patients with BRCA1/2m advanced OC after response to first-line chemotherapy in Singapore.

Effects of Vitamin D3 Supplementation on Body Composition in the VITamin D and OmegA-3 TriaL (VITAL).

CONTEXT: Although observational studies show inverse associations between vitamin D status and body weight/adiposity, there are few large randomized controlled trials (RCTs) investigating this relationship. OBJECTIVE: To determine whether vitamin D3 supplementation lowers weight or improves body composition. DESIGN: The VITamin D and OmegA-3 TriaL (VITAL) was a double-blinded, placebo-controlled RCT including 25 871 US adults. This ancillary study was completed in a sub-cohort that underwent body composition assessments at baseline and 2-year follow-up (89% retention). SETTING: Harvard Clinical and Translational Science Center in Boston. PARTICIPANTS: 771 participants (men ≥ 50 and women ≥ 55 years). INTERVENTIONS: 2 × 2 factorial design of supplemental vitamin D3 (2000 IU/day) and/or omega-3 fatty acids (1 g/day). MAIN OUTCOME MEASURES: Endpoints were 2-year changes in weight, body mass index (BMI), waist circumference, and total and/or regional fat and lean tissue measures determined by dual-energy X-ray absorptiometry. Effect modification by clinical variables and total and free 25-hydroxyvitamin D (25[OH]D) levels was explored. RESULTS: There were no effects of supplemental vitamin D3vs placebo on weight, BMI, or measures of adiposity and lean tissue. Effects did not vary by sex, race/ethnicity, fat mass index, or baseline total or free 25(OH)D levels. Vitamin D3 supplementation did slightly improve body fat percentage in participants with normal BMI at baseline, but not in the overweight or obese (P for interaction = 0.04). CONCLUSIONS: Daily vitamin D3 supplementation vs placebo in the general older population did not improve weight or body composition. Whether supplemental vitamin D3 may benefit individuals with normal BMI warrants further study.

From Emergency Department Visit to Readmission After Esophagectomy: Analysis of Burden and Risk Factors.

BACKGROUND: Frequent emergency department (ED) visits occur after esophagectomy. We aimed to identify the incidence of and risk factors for conversion from ED visit to inpatient admission. METHODS: A retrospective cohort study was performed of consecutive esophagectomies at a tertiary Canadian center (1999 to 2014). Multivariable regression analyses identified factors associated with conversion from ED visit to admission. RESULTS: There were 520 esophagectomies with 6% inhospital mortality (n = 31). Of those discharged, 29.7% (n = 145) had one or more emergency visit and 43.4% (n = 63) of these patients were readmitted to the hospital. First-time ED visits resulted in inpatient conversion 23.4% of the time (n = 34); successive ED visits resulted in increasing conversion. On multivariable analysis, anastomotic leak (adjusted odds ratio 2.45; 95% confidence interval, 1 to 6.01; P = .05) was independently associated with higher odds of conversion to admission. Sensitivity analysis using Poisson regression to model conversion as a rate identified that living in regions further away was associated with lower conversion rate to admission (risk ratio 0.35; 95% confidence interval, 0.13 to 0.94; P = .04). CONCLUSIONS: Although postesophagectomy ED utilization is high, the majority of visits do not convert to admission. With each increasing ED visit, likelihood of converting to admission increases. Anastomotic leakage was associated with higher odds of conversion to admission, possibly related to development of strictures. Access to urgent outpatient endoscopy may help reduce the incidence of ED visits and admission. Although living in regions further away is associated with lower conversion rates to admission at the index hospital, that may be due to patients utilizing closer local hospitals.

Targeted Inhibition of the E3 Ligase SCFSkp2/Cks1 Has Antitumor Activity in RB1-Deficient Human and Mouse Small-Cell Lung Cancer.

The RB1 tumor suppressor gene is mutated in highly aggressive tumors including small-cell lung cancer (SCLC), where its loss, along with TP53, is required and sufficient for tumorigenesis. While RB1-mutant cells fail to arrest at G1-S in response to cell-cycle restriction point signals, this information has not led to effective strategies to treat RB1-deficient tumors, as it is challenging to develop targeted drugs for tumors that are driven by the loss of gene function. Our group previously identified Skp2, a substrate recruiting subunit of the SCF-Skp2 E3 ubiquitin ligase, as an early repression target of pRb whose knockout blocked tumorigenesis in Rb1-deficient prostate and pituitary tumors. Here we used genetic mouse models to demonstrate that deletion of Skp2 completely blocked the formation of SCLC in Rb1/Trp53-knockout mice (RP mice). Skp2 KO caused an increased accumulation of the Skp2-degradation target p27, a cyclin-dependent kinase inhibitor, which was confirmed as the mechanism of protection by using knock-in of a mutant p27 that was unable to bind to Skp2. Building on the observed synthetic lethality between Rb1 and Skp2, we found that small molecules that bind/inhibit Skp2 have in vivo antitumor activity in mouse tumors and human patient-derived xenograft models of SCLC. Using genetic and pharmacologic approaches, antitumor activity was seen with Skp2 loss or inhibition in established SCLC primary lung tumors, in liver metastases, and in chemotherapy-resistant tumors. Our data highlight a downstream actionable target in RB1-deficient cancers, for which there are currently no targeted therapies available. SIGNIFICANCE: There are no effective therapies for SCLC. The identification of an actionable target downstream of RB1, inactivated in SCLC and other advanced tumors, could have a broad impact on its treatment.

Postoperative but not intraoperative transfusions are associated with respiratory failure after pneumonectomy.

OBJECTIVES: Transfusion of blood products has been associated with increased risk of post-pneumonectomy respiratory failure. It is unclear whether intraoperative or postoperative transfusions confer a higher risk of respiratory failure. Our objective was to assess the role of transfusions in developing post-pneumonectomy respiratory failure. METHODS: We performed a retrospective cohort study using prospectively collected data on consecutive pneumonectomies between 2005 and 2015. Patient records were reviewed for intraoperative/postoperative exposures. Univariable and multivariable analyses were performed. RESULTS: Of the 251 pneumonectomies performed during the study period, 24 (9.6%) patients suffered respiratory failure. Ninety-day mortality was 5.6% (n = 14) and was more likely in patients with respiratory failure (7/24 vs 7/227, P < 0.001). Intraoperative and postoperative transfusions occurred in 42.2% (n = 106) and 44.6% (n = 112) of patients, respectively and were predominantly red blood cells. On univariable analysis, both intraoperative (P = 0.03) and postoperative transfusion (P = 0.004) were associated with a higher risk of respiratory failure. The multivariable model significantly predicted respiratory failure with an area under curve (AUC) = 0.88 (P = 0.001). On multivariable analysis, the only independent predictors of respiratory failure were postoperative transfusions [adjusted odds ratio (aOR) 6.54, 95% confidence interval (CI) 1.74-24.59; P = 0.005] and lower preoperative forced expiratory volume (adjusted OR 0.96, 95% CI 0.93-0.99; P = 0.03). Estimated blood loss was not significantly different (P = 0.91) between those with (median 800 ml, interquartile range 300-2000 ml) and without respiratory failure (median 800 ml, interquartile range 300-2000 ml). CONCLUSIONS: Respiratory failure occurred in 9.6% of patients post-pneumonectomy and confers a higher risk of 90-day mortality. Postoperative (but not intraoperative) transfusion was the strongest independent predictor associated with respiratory failure. Intraoperative transfusion may be in reaction to active/unpredictable blood loss and may not be easily modifiable. However, postoperative transfusion may be modifiable and potentially avoidable. Transfusion thresholds should be assessed in light of potential cost-benefit trade-offs.

A phase 1 trial of SGN-CD70A in patients with CD70-positive diffuse large B cell lymphoma and mantle cell lymphoma.

Purpose This first-in-human study evaluated SGN-CD70A, an antibody-drug conjugate (ADC) directed against the integral plasma membrane protein CD70 and linked to a pyrrolobenzodiazepine (PBD) dimer, in patients with relapsed or refractory (R/R) CD70-positive non-Hodgkin lymphoma (NHL) including diffuse large B cell lymphoma (DLBCL), mantle cell lymphoma (MCL), and Grade 3b follicular lymphoma (FL3b). Methods SGN-CD70A was administered intravenously on Day 1 of 3-week cycles beginning at 8 mcg/kg with planned dose escalation to 200 mcg/kg. Due to observations of prolonged thrombocytopenia, the study was amended to dose every 6 weeks (q6wk). Results Twenty patients were enrolled and treated with SGN-CD70A. The maximum tolerated dose of SGN-CD70A was 30 mcg/kg q6wk. The most common adverse events (AEs) reported were thrombocytopenia (75%), nausea (55%), anemia (50%), and fatigue (50%). The onset for treatment-related thrombocytopenia typically occurred during Cycle 1. Most of the treatment-related events of thrombocytopenia were ≥ Grade 3. Antitumor activity in patients included 1 complete remission (CR) and 3 partial remissions (PRs), 2 of which were ongoing for at least 42.9 weeks. SGN-CD70A exposures were approximately dose proportional, with a mean terminal half-life of 3 to 5 days. Conclusions While modest single-agent activity was observed in heavily pretreated NHL patients, the applicability of SGN-CD70A is limited by the frequency and severity of thrombocytopenia, despite the long-term response with limited drug exposure.

Emergency department use is high after esophagectomy and feeding tube problems are the biggest culprit.

OBJECTIVES: Esophagectomy is a complex operation with potential for prolonged recovery. We aimed to identify the incidence of and risk factors for any and frequent emergency department visits within 1 year of esophagectomy. METHODS: A retrospective cohort study was performed looking at consecutive esophagectomies at a tertiary Canadian center (1999-2014). Multivariable analyses identified factors associated with any emergency department visits and frequent emergency department use (≥3 visits) within 1 year postesophagectomy. RESULTS: There were 520 esophagectomies with in-hospital mortality of 6% (n = 31). Of those discharged, 29.7% (n = 145) had ≥ 1 emergency department visit. Most common causes were feeding tube problems (39.3%; n = 57) and dysphagia/stricture (13.1%; n = 19). Higher income (adjusted odds ratio [aOR], 1.22; 95% confidence interval [CI], 1.04-1.42 per $10,000) and use of hybrid/minimally invasive esophagectomy (aOR, 3.24; 95% CI, 1.71-6.11) were independently associated with having emergency department visits. Patients with hybrid/minimally invasive esophagectomy were discharged earlier than others (P < .0001). Living outside of our metropolitan area (aOR, 0.36; 95% CI, 0.27-0.49) and having surgery in the later years of the study period (aOR, 0.91; 95% CI, 0.86-0.97; P = .006) were both independently associated with lower odds of emergency department visits. Forty-three patients (8.8%) were frequent emergency department users, with the most common causes of repeat emergency visits being feeding tube problems. Living outside of our metropolitan area was associated with lower odds of frequent emergency visits (aOR, 0.25; 95% CI, 0.14-0.45). CONCLUSIONS: There is high emergency department use within 1 year postesophagectomy. Patients living farther away from our hospital had a lower rate of emergency department use. It is possible that they are utilizing emergency departments nearer to home; this needs further study. Feeding tube problems are the biggest culprits and are potentially modifiable.

Surgery for acute type A aortic dissection in octogenarians is justified.

OBJECTIVE: Surgery in octogenarians with acute type A aortic dissection is commonly avoided or denied because of the high surgical morbidity and mortality reported in elderly patients. We sought to compare clinical and quality of life outcomes between octogenarians and those aged less than 80 years who underwent surgical repair at New York Medical College. METHODS: A total of 101 cases of acute type A aortic dissection repair between July 2005 and December 2011 were retrospectively analyzed, comparing 21 octogenarians with 80 concurrent patients aged less than 80 years. All patients underwent corrective surgery (ascending/hemiarch replacement in 71; Bentall in 22; David procedure in 2; Wheat procedure in 4; total arch replacement in 2) using deep hypothermic circulatory arrest. During follow-up, the RAND 36-Item Short Form Health Survey Questionnaire was used to assess quality of life. RESULTS: Octogenarians (average, 85 years; range, 80-91 years) were compared with the younger group (average, 60 years; range, 30-79 years). The 2 groups had similar preoperative characteristics, but the younger group experienced more malperfusion (40% vs 9%, P = .002), were more likely to have undergone a Bentall procedure (26% vs 5%, P = .04), and had longer circulatory arrest times (20 ± 7 minutes vs 16 ± 9 minutes, P = .03). The overall hospital mortality was 9% (9/101). Among octogenarians, there were no hospital deaths, no late deaths during follow-up (mean, 17 months; range, 1-59 months), and emotional health scores were better than those of the younger patients (P = .04). CONCLUSIONS: Surgery for acute type A aortic dissection should be offered to octogenarians because excellent surgical and quality of life outcomes can be achieved even in this elderly population.

Peripheral extracorporeal membrane oxygenation: comprehensive therapy for high-risk massive pulmonary embolism.

BACKGROUND: Although commonly reserved as a last line of defense, experienced centers have reported excellent results with pulmonary embolectomy for massive and submassive pulmonary embolism (PE). We present a contemporary surgical series for PE that demonstrates the utility of peripheral extracorporeal membrane oxygenation (pECMO) for high-risk surgical candidates. METHODS: Between June 2005 and April 2011, 29 patients were treated for massive or submassive pulmonary embolism, with surgical embolectomy performed in 26. Four high-risk patients were placed on pECMO, established by percutaneously cannulating the right atrium through a femoral vein and perfusing by a Dacron graft anastomosed to the axillary artery. A small, extracorporeal, rotary assist device was used, interposing a compact oxygenator in the circuit, and maintaining anticoagulation with heparin. RESULTS: Extracorporeal membrane oxygenation was weaned in 3 of 4 patients after 5.3 days (5, 5, and 6), with normalization of right ventricular dysfunction and pulmonary artery pressure (44.0 ± 2.0 to 24.5 ± 5.5 mm Hg) by ECHO. Follow-up computed tomographies showed several peripheral, nearly resorbed emboli in 1 case and complete resolution in 2 others. The fourth patient, not improving after 10 days, underwent surgery where an embolic liposarcoma was extracted. For all 29 cases, hospital and 30-day mortality was 0% and all patients were discharged, with average postoperative length of stay of 15 days for embolectomy and 17 days for pECMO. CONCLUSIONS: Heparin therapy with pECMO support is a rapid, effective option for patients who might benefit from pulmonary embolectomy but are at high risk for surgery.

Paradoxical response to prophylactic Didox (N-3, 4 trihydroxybenzamide) treatment in murine cytomegalovirus-infected mice.

BACKGROUND: In this study, we investigated the effect of Didox (DX) on the pathogenicity of and host responses to murine cytomegalovirus (MCMV) infection. METHODS: In vitro efficacy of DX against MCMV was determined using plaque reduction assays. For in vivo studies, mice infected with a sublethal dose (10(4) PFU) of MCMV were treated daily with DX (200 mg/kg) using either a prophylactic or delayed protocol. At predetermined intervals, target organs were removed for histopathology. Cytokine transcription and viral load were performed using real-time PCR. Serum cytokine levels were determined by ELISA, and T-cell markers by real-time PCR. RESULTS: DX (0.5-50 µM) inhibited MCMV plaque formation in vitro. However, in vivo, prophylactic DX treatment did not decrease viral load and prolonged hepatic proinflammatory cytokine transcription at days 3 and 5 post-infection, which corresponded with more severe histopathological changes observed in the liver. Significant CD8(+) T-cell marker suppression was seen, in accordance with DX-induced inhibition of lymphocyte proliferation observed in vitro. DX prolonged the recovery of MCMV-infected mice when given after infection was established. CONCLUSIONS: Despite promising MCMV inhibition in vitro, DX had no beneficial effect on MCMV disease in our model and paradoxically had adverse effects when administered prophylactically. The lack of correlation between in vitro activity and in vivo efficacy emphasizes the importance of selecting appropriate antiviral targets and of using animal models when testing new drugs.

Survival comparison of adenosquamous, squamous cell, and adenocarcinoma of the lung after lobectomy.

BACKGROUND: Primary adenosquamous carcinoma (ASC) of the lung is a rare tumor that may carry a poor prognosis. We examined a national database to see if ASC exhibited distinct clinical behavior from squamous cell (SC) and adenocarcinoma (AC) of the lung. METHODS: This is a retrospective study querying the Surveillance, Epidemiology, and End Results database to identify 872 surgical patients diagnosed with ASC, 7888 with SC, and 12,601 with AC of the lung from 1998 to 2002. Analysis characterized clinical variables to determine patterns of presentation and compared survival among the above three histologic groups after lobectomy for stage I and II disease. RESULTS: ASC represented 4.1% of the 21,361 patients examined. ASC tended toward right side (56.9%) laterality and upper lobe (60.0%) location. Compared with AC, patients with ASC and SC were more likely to be male (p < 0.0001), and ASC patients had worse histologic grade (p< 0.0001). Survival after lobectomy for stage I and II disease was significantly reduced in ASC and SC compared with AC (p < 0.0001). ASC had a significantly increased hazard ratio of 1.35 and 1.27 relative to AC and SC, respectively. Other significant negative predictors of survival included tumor grade of III and IV, stage II, age, and black ethnicity. CONCLUSIONS: This large review demonstrates that ASC is an uncommon tumor with distinct clinical behavior and worse prognosis than AC and SC. Further insight into the molecular profile of ASC is needed to determine the cause of its biologic aggressiveness.

In vivo characterization of cytokine profiles and viral load during murine cytomegalovirus-induced acute myocarditis.

BACKGROUND: Murine cytomegalovirus (MCMV) is an etiologic agent of acute and chronic myocarditis in BALB/c mice. Immunologic host responses appear to play a key role in pathogenesis but have been incompletely defined. METHODS: BALB/c mice were infected with a sublethal dose of MCMV. Cytokine transcription and viral load (measured by quantitative real-time polymerase chain reaction) and histopathological analyses were performed at specified time points. RESULTS: Increased tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and interferon (IFN)-gamma, as well as IL-10 mRNA transcripts, were detected in the hearts of infected mice starting at Day 1 post-infection (p.i.), with peak levels occurring at Day 8 p.i. (7-fold, 14-fold, 41-fold, and 16-fold higher than background, respectively). Peak cytokine transcription significantly correlated with a 10-fold increase in viral load (P<.001) at Day 8 p.i. Myocarditis-related pathological changes, measured by infiltration foci, were greatest at Day 8 p.i., corresponding with peak cytokine transcription and significantly correlated with IFN-gamma levels (P<.0001). Infiltration foci were predominantly composed of CD3(+) T cells. Cardiac calcification was observed in most infected mice predominantly over the right ventricle. Histological analysis of heart sections from mice infected with recombinant enhanced green fluorescence protein-MCMV revealed a localized and sporadic pattern of virus throughout all heart layers. CONCLUSIONS: MCMV-induced myocarditis in BALB/c mice is characterized by in vivo production of proinflammatory cytokines in a pattern correlating with MCMV viral load. The infection pattern and inflammatory response is highly localized, sporadic, and involves endocardium, epicardium, as well as the myocardium, with greatest amounts of virus detected in areas of pathologic calcification.

The diagnostic utility of cell blocks prepared from residual SurePath Pap material for detection of human papilloma virus.

This study was undertaken to determine if tissue sections from cell block prepared from the residual cellular sediment of Pap test vials could be used for immunohistochemistry (IHC) or in situ hybridization (ISH) for the detection of human papilloma virus (HPV) and to determine if these stains could clarify the nature of atypical cases in certain cases. Cases included in this retrospective study are categorized into 3 groups. Group 1 included 12 positive and 10 negative cases that were used to optimize the IHC and ISH staining protocol for the detection of HPV. Cases selected in group 2 were included to validate and verify the IHC and ISH stains. We validated 20 negative and 37 positive cases. Group 3 included 37 atypical cases. Unused material from the corresponding liquid-based (SurePath) Pap test specimens were retrieved and used to prepare paraffin-embedded cell blocks. Hematoxylin and eosin-stained cell-block sections were evaluated for abnormal cells. The IHC and ISH stain protocols for detection of HPV DNA were successfully optimized, validated, and verified. The sensitivity for the detection of HPV DNA using IHC in atypical squamous cells of undetermined significance was 80% and in atypical squamous cell, cannot rule out high grade, was 78% whereas the specificity was 100% in both lesions. Both the sensitivity and specificity for the detection of HPV DNA using the ISH were 100%. This study demonstrated that cell-block sections prepared from residual SurePath Pap test material could be used for detection of HPV DNA by both IHC and ISH and clarify the nature of atypical cells on cell-block sections.