Multicenter, Prospective Trial of Nonendoscopic Biomarker-Driven Detection of Barrett's Esophagus and Esophageal Adenocarcinoma.

INTRODUCTION: Preliminary data suggest that an encapsulated balloon (EsoCheck), coupled with a 2 methylated DNA biomarker panel (EsoGuard), detects Barrett's esophagus (BE) and esophageal adenocarcinoma (EAC) with high accuracy. The initial assay requires sample freezing upon collection. The purpose of this study was to assess a next-generation EsoCheck sampling device and EsoGuard assay in a much-enlarged multicenter study clinically enhanced by using a Clinical Laboratory Improvement Amendments of 1988-compliant assay and samples maintained at room temperature. METHODS: Cases with nondysplastic BE (NDBE), dysplastic BE (indefinite for dysplasia, low-grade dysplasia, high-grade dysplasia), EAC, junctional adenocarcinoma, plus endoscopy controls without esophageal intestinal metaplasia, were prospectively enrolled. Medical assistants at 6 institutions delivered the encapsulated balloon per orally with inflation in the stomach. The inflated balloon sampled the distal 5 cm of the esophagus and then was deflated and retracted into the capsule, preventing sample contamination. EsoGuard bisulfite sequencing assayed levels of methylated vimentin and methylated cyclin A1. RESULTS: A total of 243 evaluable patients-88 cases (median age 68 years, 78% men, 92% White) and 155 controls (median age 57 years, 41% men, 88% White)-underwent adequate EsoCheck sampling. The mean procedural time was approximately 3 minutes. Cases included 31 with NDBE, 16 with indefinite for dysplasia/low-grade dysplasia, 23 with high-grade dysplasia, and 18 with EAC/junctional adenocarcinoma. Thirty-seven NDBE and dysplastic BE cases (53%) were short-segment BE (<3 cm). Overall sensitivity was 85% (95% confidence interval 0.78-0.93) and specificity was 85% (95% confidence interval 0.79-0.90). Sensitivity for NDBE was 84%. EsoCheck/EsoGuard detected 100% of cancers (n = 18). DISCUSSION: EsoCheck/EsoGuard demonstrated high sensitivity and specificity in detecting BE and BE-related neoplasia.

Human navicular bone: a morphometric and morphological evaluation.

BACKGROUND: The composition of navicular joint complex is crucial to perform surgical interventions for multiple pathological foot aetiologies. The data on human navicular bone and its facets from Indian population remain scarce in literature. AIMS AND OBJECTIVES: To evaluate the morphometry and morphology of navicular bone. METHODOLOGY: A total of 77 (right: 40; left: 37) dried human navicular bones were used. The collected data were entered and analysed in SPSS software. RESULTS: The anteroposterior diameter of navicular bone on right side was 15.19 mm (13.92, 16.77) and on left side was 15.87 mm (13.83, 17.27). The transverse diameter on right and left sides were 34.21 mm (31.74, 36.6) and 33.59 mm (30.23, 35.43), respectively. The vertical diameter measured on the right was 22.31 mm (21.19, 23.94) and on left 22.53 mm (20.8, 24.24). Morphometric evaluation showed no significant difference between right and left navicular bones. The commonest shape for posterior facet was quadrilateral, on the right (62.5%) and left (40.5%). The most common shape of anterior facet for medial cuneiform is quadrilateral, on the right (85%) and left (89.1%). For intermediate cuneiform, triangular facet was common on the right side (72.5%) and on the left (59.5%). The lateral cuneiform facet was bean shaped on right side (72.5%) and quadrilateral on the left side (32.5%). There was a significant difference in shape distribution between right and left (P < 0.05). The median length of the groove for tibialis posterior tendon was 18.01 mm and 16.19 mm on right and left side, respectively. Cuboid facet was observed in 28 (70%) and 26 (65.9%) navicular bones on right and left sides, respectively. CONCLUSION: There is no significant difference between right and left bones with regards to morphometric parameters. Morphological evaluation revealed significant difference in the distribution of shape between right and left bones.

An unilateral double fenestration of the right external jugular vein: a rare variant.

OBJECTIVES: This report presents a rare anatomical variation, double fenestration of the External jugular vein on the right side. MATERIALS AND METHODS: During the routine dissection of a male cadaver aged 60 years, we observed a unilateral large double fenestration of the External jugular vein on the right side. RESULTS: After its formation from the posterior division of the retromandibular and posterior auricular veins, External jugular vein descended in the posterior triangle of neck. Here, it divided into medial, intermediate, and lateral veins that united again before draining into the subclavian vein. Lateral vein was the largest (7.2 cm) and intermediate and medial veins were measuring 6.4 cm each. Two large fenestrations, measuring 5.8 cm each, arranged like a "double bubble" were seen in the External jugular vein extending from fourth to sixth cervical (C4 to C6) vertebrae. The medial branch of supraclavicular nerve was seen passing superficial to the distal part of External jugular vein. On the left side, the course of External jugular vein showed a standard pattern. CONCLUSION: Surgeons must be acquainted with the varied anatomy of the superficial neck veins to prevent major bleeding during operative procedures, including carotid endarterectomy, flap operations, & central venous catheterisation.

Serum MicroRNA Signatures in Recovery From Acute and Chronic Liver Injury and Selection for Liver Transplantation.

We previously demonstrated a distinct hepatic microRNA (miRNA) signature (down-regulation of miRNA-23a, -150, - 200b, -503, and -663 and up-regulation of miRNA-20a) is associated with successful regeneration in auxiliary liver transplantation (ALT). This study aimed to evaluate whether the serum expression of this regeneration-linked miRNA signature is associated with clinical outcomes in acute and chronic liver disease. These were represented by patients with acetaminophen-induced acute liver failure (ALF; n = 18) and patients with hepatitis C virus (HCV) undergoing treatment with direct-acting antivirals (n = 56), respectively. Patients were grouped depending on their clinical outcome. Global serum miRNA expression was analyzed using polymerase chain reaction (PCR) arrays and selected miRNA expression using targeted PCR. We demonstrate that specific regeneration-linked miRNAs discriminate outcomes in both clinical scenarios. We further show that miRNA-20a, -23a, -150, -200b, -503, and -663 undergo concordant changes in expression in 3 distinct clinical settings: liver regeneration accompanying successful ALT, clinical recovery after ALF, and clinical recompensation after cure of HCV. This miRNA signature represents a potentially novel biomarker to predict outcome and optimize patient selection for liver transplantation in both acute and chronic liver disease.

Analytical performance evaluation of a commercial next generation sequencing liquid biopsy platform using plasma ctDNA, reference standards, and synthetic serial dilution samples derived from normal plasma.

BACKGROUND: Circulating tumor (ct) DNA assays performed in clinical laboratories provide tumor biomarker testing support for biopharmaceutical clinical trials. Yet it is neither practical nor economically feasible for many of these clinical laboratories to internally develop their own liquid biopsy assay. Commercially available ctDNA kits are a potential solution for laboratories seeking to incorporate liquid biopsy into their test menus. However, the scarcity of characterized patient samples and cost of purchasing validation reference standards creates a barrier to entry. In the current study, we evaluated the analytical performance of the AVENIO ctDNA liquid biopsy platform (Roche Sequencing Solutions) for use in our clinical laboratory. METHOD: Intra-laboratory performance evaluation of AVENIO ctDNA Targeted, Expanded, and Surveillance kits (Research Use Only) was performed according to College of American Pathologists (CAP) guidelines for the validation of targeted next generation sequencing assays using purchased reference standards, de-identified human plasma cell-free (cf) DNA samples, and contrived samples derived from commercially purchased normal and cancer human plasma. All samples were sequenced at read depths relevant to clinical settings using the NextSeq High Output kit (Illumina). RESULTS: At the clinically relevant read depth, Avenio ctDNA kits demonstrated 100% sensitivity in detecting single nucleotide variants (SNVs) at ≥0.5% allele frequency (AF) and 50% sensitivity in detecting SNVs at 0.1% AF using 20-40 ng sample input amount. The assay integrated seamlessly into our laboratory's NGS workflow with input DNA mass, target allele frequency (TAF), multiplexing, and number of reads optimized to support a high-throughput assay appropriate for biopharmaceutical trials. CONCLUSIONS: Our study demonstrates that AVENIO ctDNA liquid biopsy platform provides a viable alternative for efficient incorporation of liquid biopsy assays into the clinical laboratory for detecting somatic alterations as low as 0.5%. Accurate detection of variants lower than 0.5% could potentially be achieved by deeper sequencing when clinically indicated and economically feasible.

Development and evaluation of a gold nanoparticle-based immunochromatographic strip test for the detection of canine parvovirus.

Canine parvovirus (CPV) is the leading viral cause of enteritis in dogs and occurs mainly in 6- to 8-week-old pups. Rapid diagnosis of CPV under field conditions is now possible due to commercially available immunochromatographic (IC) assays. However, these commercial kits are somewhat expensive because they utilize a minimum of two monoclonal antibodies (mAbs) targeting different epitopes as capture and detector antibodies. Using only a single mAb for both capture and detection purpose may reduce the sensitivity of the assay. In the present study, efforts were made to develop an economical assay that can be utilized for diagnosis of CPV under Indian conditions with a high level of confidence. Rabbit polyclonal antibodies (pAbs) generated against recombinant truncated VP2 proteins of CPV were used as capture antibodies because they can be produced economically, while a commercial anti-CPV mAb was used as the detector antibody. The detection limit of the test strip was 6.6×105 TCID50/ml, and it specifically detected CPV-2, CPV-2a and CPV-2b while displaying no cross-reactivity with other common canine enteric pathogens. The relative sensitivity/specificity of pAb based strip test was 71%/92% and 71%/100% in relation to the hemagglutination test and a commercial IC kit, respectively, with substantial agreement. In addition, two commercially available mAbs targeting different epitopes were also used for development of another IC assay, which showed sensitivity, and specificity of 82%/87% and 90%/98% in relation to the hemagglutination test and commercial kit. Hence, the present strip test based on a combination of mAb and pAb provides an acceptable alternative for onsite and cost-effective diagnosis of CPV infection.

Commentary on Cross-Cultural Perspectives on Positive Youth Development With Implications for Intervention Research.

There is a growing focus on youth positive development issues among researchers and practitioners around the world. In this special issue of Child Development, each of the international authors provides new perspectives and understanding about youth developmental assets in different cultural settings. The present commentary (a) examines some of the cross-cultural themes that emerge from the four articles by international authors in this issue with implications for positive youth development (PYD) and (b) how intervention science can benefit by incorporating a PYD approach. As evident, youth involved in contexts that provide positive resources from significant others not only were less likely to exhibit negative outcomes, but also were more likely to show evidence of positive development.

Determination of immune status in dogs against CPV-2 by recombinant protein based latex agglutination test.

Canine parvoviral enteritis is a highly contagious viral illness caused by canine parvovirus-2 (CPV-2) which affects puppies of mainly 6-20 weeks of age. Vaccination is pivotal in preventing and controlling CPV-2 infection. Determination of antibody status is a critical determinant for successful vaccination. The hemagglutination inhibition (HI) test is 'gold standard' test for quantification of antibodies specific to CPV-2, although the execution of this test is not feasible under field conditions. The present study was undertaken to develop a point of care testing to determine immune status prior to CPV-2 vaccination or to detect seroconversion in immunized dogs by latex agglutination test (LAT) using recombinant antigen. Truncated portion of VP2 protein (tVP2) of CPV-2 was selected on the basis of antigenic indices, overexpressed the recombinant protein in E. coli system and was subsequently used in development of LAT. A total of 59 serum samples obtained from vaccinated (n = 54) and healthy unvaccinated (n = 5) dogs were tested. The positivity was observed in 85% (46/54) of these dogs with varying agglutination pattern. The overall sensitivity and specificity of latex agglutination test in comparison to HI test was recorded as 90% and 88% respectively with an agreement value of 90% (CI = 95%).

Outcomes after successful direct-acting antiviral therapy for patients with chronic hepatitis C and decompensated cirrhosis.

BACKGROUND & AIMS: Direct-acting antivirals have become widely used for patients with chronic hepatitis C virus infection with decompensated cirrhosis. Virological responses are excellent and early improvements in liver function, at least in a proportion of patients, have been observed but the longer term impact of viral clearance on end-stage liver disease complications is unclear. METHODS: Prospective study of patients with decompensated cirrhosis who received 12weeks of all-oral direct-acting antivirals through the English Expanded Access Programme. Endpoints were deaths, liver transplantation, hepatocellular carcinoma, serious decompensation events, sepsis or hospitalisations, and MELD scores between start of therapy to 15months post-treatment start. An untreated cohort of patients was retrospectively studied over 6months for comparison. RESULTS: Amongst 317/406 patients who achieved sustained virological response at 24weeks post-treatment, there were 9 deaths (3%), 17 new liver cancers (5%), 39 transplantations (12%) and 52 with serious decompensations (16%), over 15months. When compared to the first six months from treatment start and to untreated patients, there was a reduction in incidence of decompensations [30/406 (7%) in months 6-15 and 72/406 (18%) in months 0-6 for treated patients vs. 73/261 (28%) in untreated patients]. There was no significant difference in liver cancer incidence (10/406 (2.5%) in months 6-15 and 17/406 (4%) in months 0-6 for treated patients vs. 11/261 (4%) in untreated patients). CONCLUSIONS: This study suggests that antiviral therapy in patients with decompensated cirrhosis led to prolonged improvement in liver function, with no evidence of paradoxical adverse impact nor increase in liver malignancy. LAY SUMMARY: This is a report of a large group of patients in England who have hepatitis C virus (HCV) infection with advanced liver disease. They have been treated with new anti-HCV drugs, which cured the infection in the majority. This study looks at their outcomes a year following treatment, in terms of deaths, cancers and other complications of advanced liver disease. We conclude that in most patients anti-HCV treatment is beneficial even in advanced liver disease.

Impact of direct acting antiviral therapy in patients with chronic hepatitis C and decompensated cirrhosis.

BACKGROUND & AIMS: All oral direct acting antivirals (DAAs) effectively treat chronic hepatitis C virus (HCV) infection, but the benefits in advanced liver disease are unclear. We compared outcomes in treated and untreated patients with decompensated cirrhosis. METHODS: Patients with HCV and decompensated cirrhosis or at risk of irreversible disease were treated in an expanded access programme (EAP) in 2014. Treatment, by clinician choice, was with sofosbuvir, ledipasvir or daclatasvir, with or without ribavirin. For functional outcome comparison, untreated patients with HCV and decompensated cirrhosis who were registered on a database 6months before treatment was available were retrospectively studied. Primary endpoint was sustained virological response 12weeks post antiviral treatment (treated cohort) and the secondary endpoint (both cohorts) was adverse outcomes (worsening in MELD score or serious adverse event) within 6months. RESULTS: 467 patients received treatment (409 decompensated cirrhosis). Viral clearance was achieved in 381 patients (81.6%) - 209 from 231 (90.5%) with genotype 1 and 132 from 192 (68.8%) with genotype 3. MELD scores improved in treated patients (mean change -0.85) but worsened in untreated patients (mean+0.75) (p<0.0001). Patients with initial serum albumin <35g/L, aged >65 or with low (<135mmol/L) baseline serum sodium concentrations were least likely to benefit from therapy. CONCLUSIONS: All oral DAAs effectively cured HCV in patients with advanced liver disease. Viral clearance was associated with improvement in liver function within 6months compared to untreated patients. The longer term impact of HCV treatment in patients with decompensated cirrhosis remains to be determined.

Portopulmonary hypertension: Still an appropriate consideration for liver transplantation?

Liver transplantation (LT) in patients with portopulmonary hypertension (PoPH) has historically resulted in unpredictable and often poor outcomes. The United Kingdom experience for the period 1992-2012 is reported in this article. A retrospective analysis of patients, preoperatively fulfilling the PoPH European Respiratory Society Task Force on Pulmonary-Hepatic Vascular Disorders diagnostic criteria was conducted across all UK LT centers. Data collection included comorbidities, use of preoperative and postoperative pharmacotherapy, patient survival, and cause of death. To enable survival stratification, PoPH was classified as mild, moderate, or severe based on mean pulmonary pressure of <35 mm Hg, 35-49 mm Hg, and ≥50 mm Hg, respectively. Of 127 patients reported to have PoPH, just 28 fulfilled the diagnostic criteria (14 mild, 9 moderate, 5 severe). Twenty (71.4%) patients were male with median age and Model for End-Stage Liver Disease of 50 years (range, 23-62 years) and 18 (range, 6-43), respectively. Twelve (42.9%) patients died within 5 years of LT. The majority of deaths (10 of 12; 83%) occurred within the first 6 months after LT, aetiologies of which included right heart failure (n = 3), progressive PoPH (n = 2), and sepsis (n = 2). Of those receiving preoperative pharmacotherapy (n = 8), 5 are currently alive and were classified as mild to moderate PoPH. Both severe PoPH patients optimized preoperatively with pharmacotherapy died within a year of LT. Development of effective vasodilatory therapies in the setting of pulmonary arterial hypertension has led to a dramatic improvement in patient survival. The available data indicate that in this era of pharmacotherapy, PoPH in isolation no longer represents a valid consideration to transplant. Liver Transplantation 22 1637-1642 2016 AASLD.

Comparative study on nutritional and sensory quality of barnyard and foxtail millet food products with traditional rice products.

Millets have the potential to contribute to food security and nutrition, but still these are underutilized crops. The present study was undertaken with a view to analyse the physico-chemical, functional and nutritional composition of foxtail millet, barnyard millet and rice and to compare the sensory quality and nutritive value of food products from foxtail and barnyard millet with rice. Analysis of physico- chemical and functional characteristics revealed that the thousand kernel weight of foxtail millet, barnyard millet and rice was 2.5, 3.0 and 18.3 g, respectively and thousand kernel volume was 1.6, 13 2.0 and 7.1 ml, respectively. The water absorption capacity of foxtail millet, barnyard millet and rice was 1.90, 1.96 and 1.98 ml/g, respectively and water solubility index was 2.8, 1.2 and 1.0 %, respectively. Viscosity was measured for foxtail millet (1650.6 cps), barnyard millet (1581 cps) and rice (1668.3 cps). Analysis of nutritional composition showed that the moisture content of foxtail millet, barnyard millet and rice was 9.35, 11.93 and 11.91 %, respectively. The total ash, crude protein, crude fat, crude fibre and carbohydrate of foxtail millet were 3.10, 10.29, 3.06, 4.25 and 69.95 %, respectively, for barnyard millet were 4.27, 6.93, 2.02, 2.98 and 71.87 %, respectively and the corresponding values for rice were 0.59, 6.19, 0.53, 0.21 and 80.58 %, respectively. The energy value for foxtail millet, barnyard millet and rice was 349, 407 and 352 Kcal, respectively. The foxtail millet contained 30.10 mg/100 g calcium and 3.73 mg/100 g iron whereas barnyard millet contained 23.16 mg/100 g calcium and 6.91 mg/100 g iron. Values of 10 mg/100 g calcium and 0.10 mg/100 g iron were observed for rice. The formulated products viz. laddu, halwa and biryani from foxtail millet, barnyard millet and rice (control) were analysed for their sensory qualities. Among the products prepared, there was non significant difference with regard to the colour, flavor, texture, appearance and overall acceptability of foxtail and barnyard millet laddu and halwa when compared to control. Foxtail millet biryani was most acceptable compared to barnyard millet and control biryani. Nutritive value of formulated products was calculated and it was compared with the rice. The protein, fat and fibre content of the formulated products from foxtail and barnyard millet were higher than the rice products. Thus from the present study it was concluded that the foxtail millet and barnyard millet are superior in nutritive value to rice and have potential for use in traditional food products.

Unique variation in the course of maxillary artery in infratemporal fossa: a case report.

Maxillary artery is one of the key contents of the infratemporal fossa. Mandibular nerve and its branches form a clinically important relation of maxillary artery in this region. A comprehensive knowledge of variations of maxillary artery in the fossa is of special relevance in oral maxillofacial surgeries, management of epistaxis, intractable neuralgias or headaches. We found a unique variation of maxillary artery, presenting bilaterally, in relation to branches of mandibular nerve. During routine dissection in a 55-year-old male cadaver, maxillary artery was seen passing deep to lateral pterygoid muscle and crossed through the nerve loop formed between two roots of auriculotemporal nerve and posterior division of mandibular nerve. Further course of maxillary artery was medial to the posterior division of mandibular nerve. Maxillary artery gave its middle meningeal artery branch as it traversed through the nerve loop. A tortuous course taken by maxillary artery can lead to its entrapment causing headaches or nerve irritation presenting with neuralgia.

Large ovarian mature teratoma with gliomatosis peritonei in a young female: A case report.

Gliomatosis peritonei is an extremely rare condition usually associated with either immature teratoma or, less commonly, mature teratoma. We present a case of a young female with long-standing progressive abdominal distension, who was diagnosed with mature ovarian teratoma with gliomatosis peritonei and gross ascites. The final diagnosis in this case was determined through the correlation of imaging, operative, and histopathological findings. The presence of enhancing peritoneal nodules usually leads to a suspicion of peritoneal carcinomatosis or abdominal tuberculosis, especially in endemic regions; however, gliomatosis peritonei should always be considered in the differential diagnosis, particularly when associated with teratomas. Radiological findings combined with histopathological reports are valuable in reaching the final diagnosis in these cases.

Analytical Validation of a DNA Methylation Biomarker Test for the Diagnosis of Barrett's Esophagus and Esophageal Adenocarcinoma from Samples Collected Using EsoCheck®, a Non-Endoscopic Esophageal Cell Collection Device.

Barrett's esophagus (BE) is a known precursor to esophageal adenocarcinoma (EAC). Guidelines recommend BE screening in populations with multiple risk factors, for which non-endoscopic esophageal cell collection with biomarker testing is considered as an acceptable alternative to esophagogastroduodenoscopy (EGD). The aim of this study was to evaluate analytical performance characteristics of EsoGuard® (EG), a DNA methylation biomarker assay, as a laboratory-developed test (LDT) in esophageal samples collected with the swallowable EsoCheck® (EC) device. EG is a next-generation sequencing (NGS) assay that evaluates methylated vimentin (VIM) and cyclin A1 (CCNA1), clinically validated biomarkers for the detection of BE and EAC. The studies were conducted according to standards of College of American Pathology (CAP), Clinical Laboratory Improvement Amendments (CLIA), and New York (NY) state requirements for the analytical validation of molecular assays. Comparison to Sanger sequencing showed that EG was 100% accurate at all 31 CpG sites evaluated by the assay. The analytical sensitivity, specificity, and accuracy of the assay were 89%, 100%, and 96%, respectively. Intra- and inter-assay precision was 100%. The limit of detection (LOD) was 1 in 400 methylated cells, and the reference range was 84%. In summary, EsoGuard demonstrates high analytical accuracy, repeatability, and reproducibility in samples collected using the EsoCheck device.

The estrogen-responsive Agr2 gene regulates mammary epithelial proliferation and facilitates lobuloalveolar development.

Agr2 is a putative protein disulfide isomerase (PDI) initially identified as an estrogen-responsive gene in breast cancer cell lines. While Agr2 expression in breast cancer is positively correlated with estrogen receptor (ER) expression, it is upregulated in both hormone dependent and independent carcinomas. Several in vitro and xenograft studies have implicated Agr2 in different oncogenic features of breast cancer; however, the physiological role of Agr2 in normal mammary gland development remains to be defined. Agr2 expression is developmentally regulated in the mammary gland, with maximum expression during late pregnancy and lactation. Using a mammary gland specific knockout mouse model, we show that Agr2 facilitates normal lobuloalveolar development by regulating mammary epithelial cell proliferation; we found no effects on apoptosis in Agr2(-/-) mammary epithelial cells. Consequently, mammary glands of Agr2(-/-) females exhibit reduced expression of milk proteins, and by two weeks post-partum their pups are smaller in size. Utilizing a conditional mouse model, we show that Agr2 constitutive expression drives precocious lobuloalveolar development and increased milk protein expression in the virgin mammary gland. In vitro studies using knock down and overexpression strategies in estrogen receptor positive and negative mammary epithelial cell lines demonstrate a role for Agr2 in estradiol-induced cell proliferation. In conclusion, the estrogen-responsive Agr2, a candidate breast cancer oncogene, regulates epithelial cell proliferation and lobuloalveolar development in the mammary gland. The pro-proliferative effects of Agr2 may explain its actions in early tumorigenesis.

Hepatocyte senescence predicts progression in non-alcohol-related fatty liver disease.

BACKGROUND & AIMS: Models of non-alcohol-related fatty liver disease (NAFLD) reveal features of accelerated ageing, such as impaired regeneration, and an increased risk of hepatocellular carcinoma. The relation between accelerated ageing, disease progression and clinical outcome has not been previously investigated and is the subject of the current study. METHODS: Liver sections from 70 patients with NAFLD (105 biopsies) and 60 controls were studied for telomere length, nuclear area, DNA damage and cell cycle phase markers, using quantitative fluorescent in situ hybridization and immunohistochemistry. RESULTS: Hepatocyte telomeres were shorter in NAFLD than controls (p <0.0001). Hepatocytes in NAFLD demonstrated lack of cell cycle progression beyond G1/S phase and high-level expression of p21, the universal cell cycle inhibitor (p=0.001). γ-H(2)AX expression increased with steatosis (p=0.01), indicating DNA damage, and was associated with shorter hepatocyte telomeres (p <0.0001). Hepatocyte p21 expression correlated with fibrosis stage and diabetes mellitus, independently (p <0.001 and p=0.002, respectively). Further analysis revealed that an adverse liver-related outcome was strongly associated with higher hepatocyte p21 expression and greater hepatocyte nuclear area (p=0.02 and p=0.006), but not with telomere length. In paired biopsies, changes in hepatocyte p21 expression and nuclear area mirrored changes in fibrosis stage (p=0.01 and p=0.006, respectively). CONCLUSIONS: These findings are consistent with hepatocyte senescence and permanent cell cycle arrest in NAFLD. Hepatocyte senescence correlated closely with fibrosis stage, diabetes mellitus, and clinical outcome. Hepatocyte p21 expression could be used as a prognostic marker and for stratification in clinical studies.

Sustained telomere length in hepatocytes and cholangiocytes with increasing age in normal liver.

UNLABELLED: Telomeres, a validated biomarker of aging, comprise multiple nucleotide repeats capping chromosomes that shorten with each cell cycle until a critical length is achieved, precipitating cell senescence. Only two previous studies focused on the effect of aging in "normal" liver tissue, but these studies were compromised by small sample size, limited age range, tissue derived from individuals with an increased risk of senescence, and the use of liver homogenates. We developed a robust large-volume, four-color quantitative fluorescent in situ hybridization technique to measure telomere length in large numbers of hepatocytes, Kupffer cells, hepatic stellate cells, CD4-positive and CD8-positive lymphocytes, and cholangiocytes. Following validation against the gold standard (Southern blotting), the technique was applied to normal archived paraffin-embedded liver tissue obtained following reperfusion of implanted donor liver. We studied 73 highly selected donors aged 5-79 years with a short medical illness preceding death and no history of liver disease, reperfusion injury, or steatosis and normal graft function 1-year posttransplantation. Cholangiocytes had significantly longer telomeres compared with all other intrahepatic lineages over a wide age range (P < 0.05). Age-related telomere attrition was restricted to sinusoidal cells (i.e., Kupffer cells [P = 0.0054] and stellate cells [P = 0.0001]). Cholangiocytes and hepatocytes showed no age-related telomere shortening. CONCLUSION: In normal liver and over a broad age range, cholangiocytes have longer telomeres than all other intrahepatic lineages. Age-related telomere length decline is restricted to Kupffer cells and stellate cells.

Morphology of middle cerebral artery using computed tomography angiographic study in a tertiary care hospital.

Increased tortuosity of vessel is associated with high incidence of plaque formation leading to atherosclerosis. Surgical procedures are done after analyzing morphology of middle cerebral artery (MCA). However, literature describing MCA morphology using computed tomography angiography (CTA) is limited, so this study was planned to determine its incidence in Indian population. Datasets of CTA from 289 patients (180 males and 109 females), average age: 49.29±16.16 years (range: 11 to 85 years), from a tertiary care hospital were systematically reviewed for morphology of MCA. Cases involving aneurysms and infarcts were excluded. Four shapes of MCA were recognized: straight, U, inverted U, and S-shaped. MCA was straight in 44% (254/578), U-shaped in 37% (215/578), S shaped in 15% (89/578) and inverted U-shaped in 3% (20/578) cases. In males, MCA was straight in 46% (166/360), U-shaped in 37% (134/360), S-shaped in 16% (58/360) and inverted U-shaped in 4% (14/360) cases. In females, MCA was straight in 42% cases (92/218), U-shaped in 37% (81/218), S-shaped in 17% (36/218) and inverted U-shaped in 4% (9/218). On comparing shape with various age groups using chi square test, U shaped (P≤0.001) and S-shaped (P=0.003) MCA were found to be statistically significant. The incidence of straight shape was higher in advanced age group (>60 years). Knowledge of MCA shape will be useful for clinicians and surgeons in successful endovascular recanalization. Also, this data would help surgeons during neurointerventional procedures.