bluuebunny/crossref_metadata_2025 huggingface dataset:
Predictors and Health Consequences of Epigenetic Changes Associated with Excess Body Weight in Women of Child-bearing Age
Authors: Piyathilake C , Badiga S , Johanning G , Alvarez R , Partridge E
Published: March 2011

Background: Epigenetic alterations occurring during pregnancy have recently emerged as important factors for developmental programming of the fetus leading to obesity-related diseases in children. However, the role of excess body weight (EBW) in the modification of epigenetic patterns or its health consequences during child-bearing age is largely unknown. Because a lower degree of DNA methylation of long interspersed nucleotide element-1 (LINE-1) in PBMCs was shown to be associated with a higher risk of developing obesity-related diseases, for example cancer, the purpose of this study was to (1) evaluate the influence of indicators of obesity (BMI, WC, and % body fat) on PBMC LINE-1 methylation, (2) determine the predictors of PBMC LINE-1 methylation, and (3) determine the influence of PBMC LINE-1 methylation on biomarkers of obesity-related diseases.

Methods: The study population consisted of 470 child-bearing age women. We quantified the degree of PBMC LINE-1 methylation by pyrosequencing. Folate concentrations were measured using a microbiological assay. The degree of LINE-1 methylation (> median vs. ≤ median) was the dependent variable in logistic models that specified BMI (>25 vs. ≤ 25), WC (>88 cm vs. ≤ 88 cm), or % body fat (>33% vs. ≤ 33%) separately as the independent predictors of primary interest, adjusting for other relevant variables. The predictors and determinants of lower LINE-1 methylation were evaluated among women with EBW.

Results: Women with higher BMI, WC, or % body fat were 2.0, 1.9, and 1.8 times more likely to have lower LINE-1 methylation, respectively (P = 0.003, 0.005, and 0.01). The predictors and determinants of lower LINE-1 methylation yielded similar patterns with all three indicators of obesity. The following results are based on models run with BMI as the indicator for EBW. Women with higher plasma folate concentrations were less likely to have lower LINE-1 methylation (OR = 0.54, P = 0.0009). Higher LINE-1 methylation was associated with lower insulin resistance as indicated by HOMA (OR = 0.50, P = 0.02).

Conclusions: EBW-associated lower LINE-1 methylation in women of child-bearing age appears to have significant, and potentially transgenerational, health consequences. Higher folate status may exert beneficial effects on obesity-related health outcomes.
Anaplastic and Poorly Differentiated Thyroid Carcinoma: Therapeutic Strategies and Treatment Outcome of 52 Consecutive Patients
Authors: Siironen P. , Hagström J. , Mäenpää H.O. , Louhimo J. , Heikkilä A. , Heiskanen I. , Arola J. , Haglund C.
Published: March 2011

Anaplastic thyroid carcinoma (ATC) is one of the most lethal malignancies; poorly differentiated thyroid carcinoma (PDTC) is a new diagnosis for rare aggressive thyroid tumours. Surgery is often considered the only chance for survival, but the benefit of surgery and subsequent multimodal therapy is unclear. We retrospectively analyzed the outcome of 44 ATC and 8 PDTC consecutive patients treated at Helsinki University Central Hospital between 1990 and 2008. All ATC and PDTC cases were re-examined and reclassified histologically. Median survival was only 3.1 months for ATC, but 3.7 years for PDTC. Most patients in both groups eventually died of cancer. ATC patients were older than PDTC patients (74 vs. 66 years). Nodal and distant metastases had a negative impact on survival (ATC; p = 0.038, p = 0.008). Long-term survivors in both groups were stage N0M0 at presentation. Multimodal therapy was successful for 9 (20%) ATC patients, and their median survival was the longest (11.6 months) among treatment groups. Most PDTC patients (88%) underwent total thyroidectomy followed by radioiodine ablation; the only 2 who received chemotherapy survived longest. Although ATC and PDTC are both aggressive thyroid carcinomas, multimodal therapy for both can provide a chance of prolonged survival in patients with locoregional disease.
Cefepime Resistance and Associated Risk Factors among Escherichia coli Strains Isolated from Clinical Specimens
Authors: Mansouri Majid , Ramazanzadeh Rashid , Norabadi Parisa
Published: March 2011

Cefepime is active against bacteria producing chromosomally and plasmid-mediated extended broad-spectrum β-lactamase enzymes. The aim of this study was to evaluate risk factors for acquisition of cefepime resistance in Escherichia coli strains among hospitalized patients in a university hospital in Sanandaj, Iran. The study was a case-control investigation. A case patient was defined as a patient who had one isolate of a cefepime-resistant E. coli strain. A control patient was defined as a patient who had one isolate of a cefepime-sensitive E. coli strain. Cefepime resistance was determined by HiComb MIC tests (HIMEDIA, India). Out of the 255 total isolates, 73 (28.6%) were resistant to cefepime. The previous treatment of cefepime was a risk factor for acquisition of a cefepime-resistant isolate (OR = 6.32, 95% CI: 1.5–25.19, p < 0.007). The use of a ventilator was considered to be a risk for acquisition of a cefepime-resistant isolate (OR = 6.25, 95% CI: 1.86–21.02, p <0.002). The use of a catheter was also found to be a risk factor for acquisition of cefepime resistance (OR = 6.28, 95% CI: 1.86–21.02, p <0.001). There was a significant correlation between days of stay in hospital wards and cefepime resistance (p < 0.003). The main risk factors associated with cefepime resistance were previous treatment with cefepime, use of ventilator, use of catheter and days of stay in ward. More studies are needed to evaluate the role of these factors in order to control the spread of drug resistance.
Kurashiki Prehospital Stroke Subtyping Score (KP3S) as a Means of Distinguishing Ischemic from Hemorrhagic Stroke in Emergency Medical Services
Authors: Yamashita Shinji , Kimura Kazumi , Iguchi Yasuyuki , Shibazaki Kensaku , Watanabe Masao , Iwanaga Takeshi
Published: March 2011

Background and Purpose: The aim of this study was to devise a new ischemic stroke (IS)/hemorrhagic stroke (HS) stroke score to distinguish IS from HS for emergency medical services (EMS). Materials and Methods: We studied patients with IS and HS who were admitted within 6 h of onset and transferred by the EMS. We compared characteristics of IS and HS to devise a new IS/HS score. Results: A total of 227 patients (median age, 71 years; 139 males; IS, 127 patients; HS, 100 patients) were included in the study. On multivariate analysis following univariate analysis, presence of atrial fibrillation, diastolic blood pressure <100 mm Hg and lack of disturbance of consciousness were independently associated with IS. The following score was devised to distinguish IS from HS in EMS: Kurashiki Prehospital Stroke Subtyping Score (KP3S) = (presence of atrial fibrillation) · 2 + (diastolic blood pressure <100 mm Hg) + (lack of disturbance of consciousness). When KP3S was greater than 1, sensitivity for IS was 64% and specificity 85%. The C statistic of KP3S was 0.805. Conclusion: KP3S is useful for distinguishing IS from HS and for the evaluation of stroke patients by EMS.
Clouded Leopard Predation on a Wild Juvenile Siamang
Authors: Morino Luca
Published: March 2011

I report the first documented case of a wild siamang killed by a clouded leopard. The event, involving a juvenile male (weighing 3.7 kg), occurred at dawn, near a known sleeping tree in a lowland tropical forest in southern Sumatra (Indonesia). Examination of the siamang’s body revealed that the first 2 cervical vertebrae were crushed and the face was partially eaten. This attack highlights the potential importance of predation as an evolutionary pressure for hylobatids, and poses some questions about the antipredator adaptations of this population of siamang.
Piglet Model for Studying Esophageal Regrowth after Resection and Interposition of a Silicone Stented Small Intestinal Submucosa Tube
Authors: Jönsson L. , Gatzinsky V. , Jennische E. , Johansson C. , Nannmark U. , Friberg L.G.
Published: March 2011

Purpose: To investigate the use and subsequent healing of a silicone stented small intestinal submucosa (SIS) tube as a full-circumference replacement following surgical resection of the esophagus in piglets. Material and Methods: Three centimeters of the intrathoracic esophagus was replaced with a silicone stented SIS tube (Cook Medical) in 6 growing piglets. The esophageal stent was retained for 4 weeks. Esophageal dilations were performed, if needed, after stent removal. Results: The piglets were sacrificed 1–17 weeks after surgery. Recurrent dilations were needed after stent removal. Histology showed that the gap between the resection margins was filled with new loose connective tissue consisting of fibroblasts and few inflammatory cells. In this tissue, intense angiogenesis was seen at the early time points, which then gave way to the proliferation of immature-looking smooth-muscle-like cells in the submucosa, which appeared to stem from the pericytes of the ingrowing capillaries. Conclusions: Through using a stented SIS tube as a circumferential esophageal replacement in a piglet model, this study suggests that pericytes from ingrowing capillaries may play a role in the remodeling of the SIS mesh. It remains to be seen if this process gives a favorable end result because stricture formation after stent removal remains a problem.
Factors Associated with Medication Adherence in Patients with Chronic Obstructive Pulmonary Disease
Authors: Agh Tamas , Inotai Andras , Meszaros Agnes
Published: April 2011

Background: Predictors of medication adherence are not well known in chronic obstructive pulmonary disease (COPD). It is therefore necessary to identify factors associated with adherence to improve the effectiveness of COPD management within real-world situations. Objectives: The goals of this study were to estimate adherence to respiratory medication and to identify factors related to adherence in COPD patients. Methods: This was an observational, cross-sectional study conducted on a sample of COPD outpatients. The following information was obtained: adherence to respiratory therapy (Morisky Medication Adherence Scale), age, gender, smoking status, COPD severity [Global Initiative for Chronic Obstructive Lung Disease (GOLD) stage], lung function [post-bronchodilator forced expiratory volume in 1 s (FEV1)], treatment regimen for COPD, COPD medication costs per month paid by the patient and health-related quality of life (EuroQol 5-dimension questionnaire). A multivariate logistic regression analysis was performed to identify the independent predictors of adherence. Results: Of the 170 participants (mean age 63.8 years, 41.8% male), 58.2% reported optimal adherence. Adherence to respiratory therapy was associated with age, current smoking status, number of respiratory drugs, number of daily respiratory drug doses and quality of life (p < 0.005). Adherence to respiratory therapy was not related to gender, GOLD stage, FEV1 or COPD medication costs. Conclusions: Adherence to COPD medication regimens is poor. Less frequent dosing regimens could be an effective method to enhance adherence to respiratory therapy. Quality-of-life monitoring within clinical practice settings could facilitate improved medication adherence.
Influence of Body Mass Index on Growth Hormone Responses to Classic Provocative Tests in Children with Short Stature
Authors: Lee Hae Sang , Hwang Jin Soon
Published: April 2011

Background: The diagnosis of growth hormone (GH) deficiency is based on a reduced peak GH response to provocative tests. However, the provocative tests are poorly reproducible and GH secretion is regulated by physiological parameters, such as body weight and puberty. The aim of this study was to assess the influence of body mass index (BMI) on GH response to provocative testing and to analyze the reproducibility of GH stimulation test. Methods: Clinical data were collected retrospectively by chart review from the Pediatric Endocrine Unit at the Ajou University Hospital. A total of 187 subjects with short stature who completed a GH stimulation testing between 2003 and 2009 were included in the study. Results: Of the 187 subjects, 66 (35.3%) had GH deficiency, while 121 (64.7%) were categorized as having idiopathic short stature. Reliability was calculated for 48 patients with idiopathic short stature who underwent the GH stimulation test twice. A GH response ≧10 ng/ml after retesting was found in 39 patients (81.3%) and a GH response <10 ng/ml was found in 9 patients (18.7%). In a stepwise multivariate analysis, BMI was a significantly independent predictor of peak GH. Elevated BMI was negatively associated with peak plasma GH levels. Conclusions: The lack of reliability of GH values in response to pharmacological stimuli should be taken into account in the diagnosis of GH deficiency. Also, higher BMI is associated with lower GH secretion. BMI should be measured and GH results appropriately interpreted for all subjects undergoing GH stimulation testing.
Runx2-Upregulated Receptor Activator of Nuclear Factor κB Ligand in Calcifying Smooth Muscle Cells Promotes Migration and Osteoclastic Differentiation of Macrophages
Authors: Byon Chang Hyun , Sun Yong , Chen Jianfeng , Yuan Kaiyu , Mao Xia , Heath Jack M. , Anderson Peter G. , Tintut Yin , Demer Linda L. , Wang Deli , Chen Yabing
Published: April 2011

Objective—

Clinical and experimental studies demonstrate the important roles of vascular smooth muscle cells (VSMC) in the pathogenesis of atherosclerosis. We have previously determined that the osteogenic transcription factor Runx2 is essential for VSMC calcification. The present study characterized Runx2-regulated signals and their potential roles in vascular calcification.

Methods and Results—

In vivo studies with atherogenic apolipoprotein E
−/−
mice demonstrated that increased oxidative stress was associated with upregulation of Runx2 and receptor activator of nuclear factor κB ligand (RANKL), which colocalized in the calcified atherosclerotic lesions and were juxtaposed to infiltrated macrophages and osteoclast-like cells that are positively stained for an osteoclast marker, tartrate-resistant acid phosphatase. Mechanistic studies using RNA interference, a luciferase reporter system, chromatin immunoprecipitation, and electrophoretic mobility shift assays indicated that Runx2 regulated the expression of RANKL via a direct binding to the 5′-flanking region of the RANKL. Functional characterization revealed that RANKL did not induce VSMC calcification, nor was RANKL required for oxidative stress–induced VSMC calcification. Using a coculture system, we demonstrated that VSMC-expressed RANKL induced migration as well as differentiation of bone marrow-derived macrophages into multinucleated, tartrate-resistant acid phosphatase–positive osteoclast-like cells. These effects were inhibited by the RANKL antagonist osteoprotegerin and with VSMC deficient in Runx2 or RANKL.

Conclusion—

We demonstrate that Runx2 directly binds to the promoter and controls the expression of RANKL, which mediates the crosstalk between calcifying VSMC and migration and differentiation of macrophages into osteoclast-like cells in the atherosclerotic lesions. Our studies provide novel mechanistic insights into the regulation and function of VSMC-derived RANKL in the pathogenesis of atherosclerosis and vascular calcification.
Signal Evolution and Infarction Risk for Apparent Diffusion Coefficient Lesions in Acute Ischemic Stroke Are Both Time- and Perfusion-Dependent
Authors: An Hongyu , Ford Andria L. , Vo Katie , Powers William J. , Lee Jin-Moo , Lin Weili
Published: April 2011

Background and Purpose—

This study aimed to examine the temporal relationship between tissue perfusion and apparent diffusion coefficient (ADC) changes within 6 hours of ischemic stroke onset and how different reperfusion patterns may affect tissue outcome in ADC lesions.

Methods—

Thirty-one participants were sequentially imaged at 3 hours, 6 hours, and 1 month post-stroke. Three regions of interest (ROIs) were defined within initial ADC lesions: ROI (1)
reperf_3hour
hyperacute reperfusion (within 3 hours), ROI (2)
reperf_6hour
acute reperfusion (3 to 6 hours), and ROI (3)
nonreperf
no reperfusion (by 6 hours). For each ROI, changes in ADC (ΔADC) from 3 to 6 hours and risks of infarction were examined.

Results—

The magnitude of initial ADC reduction was similar in all 3 ROIs (
P
=0.51). ΔADC was strongly associated with reperfusion (
P
<0.0001) but not with initial ADC reduction (
P
=0.83). ΔADC in ROI (1)
reperf_3hour
and ROI (2)
reperf_6hour
was significantly larger than that of ROI (3)
nonreperf
(
P
<0.05). Positive ΔADC was obtained from 3 to 6 hours in ROI (1)
reperf_3hour
that had restored perfusion before 3 hours, demonstrating a temporal delay between reperfusion and ADC changes. Risks of infarction were significantly higher in ROI (3)
nonreperf
than those in ROI (1)
reperf_3hour
and ROI (2)
reperf_6hour
.

Conclusions—

Improvement in ADC did not occur coincidently with reperfusion but showed a temporal delay. Regions with similar initial ADC reductions at 3 hours had different evolution of ADC and infarction risks depending on when or if tissue reperfused. These findings provide a physiological basis for the observation that a single ADC measurement at a fixed time after stroke onset may not accurately predict tissue outcome.
I Did It: Unconscious Expectation of Sensory Consequences Modulates the Experience of Self-agency and Its Functional Signature
Authors: Gentsch Antje , Schütz-Bosbach Simone
Published: March 2011

The ability to recognize oneself in voluntary action is called the sense of agency and refers to the experience of causing one's own actions and their sensory consequences. This form of self-awareness is important not only for motor control but also for social interactions and the ascription of causal responsibility. Here, we examined the sense of agency at early and prereflective stages of action perception using ERPs. Subjects performed a visual forced-choice response task in which action effects were either caused by the subject or by the computer. In addition, to modulate the conscious experience of agency, action effects were subliminally primed by the presentation of congruent, incongruent, or neutral effect stimuli before the action. First, we observed sensorimotor attenuation in the visual ERP selectively for self-generated action effects. That is, the N1 component, a negative deflection around 100 msec after a visual stimulus, was smaller in amplitude for visual effects caused by the subject as compared with effects caused by the computer. Second, congruent effect priming enhanced the explicit judgment of agency and further reduced the N1 amplitude for self-generated effects, although effect primes were not consciously processed. Taken together, these results provide evidence of a top–down modulation of sensory processing of action effects by prior effect information and support the neurophysiological mechanism of sensorimotor attenuation as underlying self-registration in action. Our findings suggest that both efferent information and prior thoughts about the action consequence provide important cues for a prereflective form of the experience of being an agent.
Is Morality Unified? Evidence that Distinct Neural Systems Underlie Moral Judgments of Harm, Dishonesty, and Disgust
Authors: Parkinson Carolyn , Sinnott-Armstrong Walter , Koralus Philipp E. , Mendelovici Angela , McGeer Victoria , Wheatley Thalia
Published: March 2011

Much recent research has sought to uncover the neural basis of moral judgment. However, it has remained unclear whether “moral judgments” are sufficiently homogenous to be studied scientifically as a unified category. We tested this assumption by using fMRI to examine the neural correlates of moral judgments within three moral areas: (physical) harm, dishonesty, and (sexual) disgust. We found that the judgment of moral wrongness was subserved by distinct neural systems for each of the different moral areas and that these differences were much more robust than differences in wrongness judgments within a moral area. Dishonest, disgusting, and harmful moral transgression recruited networks of brain regions associated with mentalizing, affective processing, and action understanding, respectively. Dorsal medial pFC was the only region activated by all scenarios judged to be morally wrong in comparison with neutral scenarios. However, this region was also activated by dishonest and harmful scenarios judged not to be morally wrong, suggestive of a domain-general role that is neither peculiar to nor predictive of moral decisions. These results suggest that moral judgment is not a wholly unified faculty in the human brain, but rather, instantiated in dissociable neural systems that are engaged differentially depending on the type of transgression being judged.
Involvement of the Primary Motor Cortex in Controlling Movements Executed with the Ipsilateral Hand Differs between Left- and Right-handers
Authors: van den Berg Femke E. , Swinnen Stephan P. , Wenderoth Nicole
Published: March 2011

Unimanual motor tasks, specifically movements that are complex or require high forces, activate not only the contralateral primary motor cortex (M1) but evoke also ipsilateral M1 activity. This involvement of ipsilateral M1 is asymmetric, such that the left M1 is more involved in motor control with the left hand than the right M1 in movements with the right hand. This suggests that the left hemisphere is specialized for movement control of either hand, although previous experiments tested mostly right-handed participants. In contrast, research on hemispheric asymmetries of ipsilateral M1 involvement in left-handed participants is relatively scarce. In the present study, left- and right-handed participants performed complex unimanual movements, whereas TMS was used to disrupt the activity of ipsilateral M1 in accordance with a “virtual lesion” approach. For right-handed participants, more disruptions were induced when TMS was applied over the dominant (left) M1. For left-handed participants, two subgroups could be distinguished, such that one group showed more disruptions when TMS was applied over the nondominant (left) M1, whereas the other subgroup showed more disruptions when the dominant (right) M1 was stimulated. This indicates that functional asymmetries of M1 involvement during ipsilateral movements are influenced by both hand dominance as well as left hemisphere specialization. We propose that the functional asymmetries in ipsilateral M1 involvement during unimanual movements are primarily attributable to asymmetries in the higher-order areas, although the contribution of transcallosal pathways and ipsilateral projections cannot be completely ruled out.
Mechanisms Underlying the Emergence of Object Representations during Infancy
Authors: Scott Lisa S.
Published: March 2011

The effects of individual versus category training, using behavioral indices of stimulus discrimination and neural ERPs indices of holistic processing, were examined in infants. Following pretraining assessments at 6 months, infants were sent home with training books of objects for 3 months. One group of infants was trained with six different strollers labeled individually, and another group was trained with the same six strollers labeled at the category level (i.e., “stroller”). Infants returned for posttraining assessments at 9 months. Discrimination of objects was facilitated for infants trained with the individually labeled strollers but was unchanged after training at the category level. Relative to pretraining and to category-level training, individual-level training resulted in increased holistic processing of strollers recorded over occipital brain regions. These results suggest that labeling nonface objects individually, in infancy, facilitates discrimination and leads to the emergence of holistic neural representations not present with category-level labeling.
Predictive Coding and Pitch Processing in the Auditory Cortex
Authors: Kumar Sukhbinder , Sedley William , Nourski Kirill V. , Kawasaki Hiroto , Oya Hiroyuki , Patterson Roy D. , Howard Matthew A. , Friston Karl J. , Griffiths Timothy D.
Published: March 2011

In this work, we show that electrophysiological responses during pitch perception are best explained by distributed activity in a hierarchy of cortical sources and, crucially, that the effective connectivity between these sources is modulated with pitch strength. Local field potentials were recorded in two subjects from primary auditory cortex and adjacent auditory cortical areas along the axis of Heschl's gyrus (HG) while they listened to stimuli of varying pitch strength. Dynamic causal modeling was used to compare system architectures that might explain the recorded activity. The data show that representation of pitch requires an interaction between nonprimary and primary auditory cortex along HG that is consistent with the principle of predictive coding.
Choosing Our Words: Retrieval and Selection Processes Recruit Shared Neural Substrates in Left Ventrolateral Prefrontal Cortex
Authors: Snyder Hannah R. , Banich Marie T. , Munakata Yuko
Published: March 2011

When we speak, we constantly retrieve and select words for production in the face of multiple possible alternatives. Our ability to respond in such underdetermined situations is supported by left ventrolateral prefrontal cortical (VLPFC) regions, but there is active debate about whether these regions support (1) selection between competing alternatives, (2) controlled retrieval from semantic memory, or (3) selection and controlled retrieval in distinct subregions of VLPFC (selection in mid-VLPFC and controlled retrieval in anterior VLPFC). Each of these theories has been supported by some prior evidence but challenged by other findings, leaving the debate unresolved. We propose that these discrepancies in the previous literature reflect problems in the way that selection and controlled retrieval processes have been operationalized and measured. Using improved measures, we find that shared neural substrates in left VLPFC support both selection and controlled retrieval, with no dissociation between mid and anterior regions. Moreover, selection and retrieval demands interact in left VLPFC, such that selection effects are greatest when retrieval demands are low, consistent with prior behavioral findings. These findings enable a synthesis and reinterpretation of prior evidence and suggest that the ability to respond in underdetermined situations is affected by both selection and retrieval mechanisms for verbal material subserved by left VLPFC, and these processes interact in meaningful ways.
Transient Perceptual Neglect: Visual Working Memory Load Affects Conscious Object Processing
Authors: Emrich Stephen M. , Burianová Hana , Ferber Susanne
Published: March 2011

Visual working memory (VWM) is a capacity-limited cognitive resource that plays an important role in complex cognitive behaviors. Recent studies indicate that regions subserving VWM may play a role in the perception and recognition of visual objects, suggesting that conscious object perception may depend on the same cognitive and neural architecture that supports the maintenance of visual object information. In the present study, we examined this question by testing object processing under a concurrent VWM load. Under a high VWM load, recognition was impaired for objects presented in the left visual field, in particular when two objects were presented simultaneously. Multivariate fMRI revealed that two independent but partially overlapping networks of brain regions contribute to object recognition. The first network consisted of regions involved in VWM encoding and maintenance. Importantly, these regions were also sensitive to object load. The second network comprised regions of the ventral temporal lobes traditionally associated with object recognition. Importantly, activation in both networks predicted object recognition performance. These results indicate that information processing in regions that mediate VWM may be critical to conscious visual perception. Moreover, the observation of a hemifield asymmetry in object recognition performance has important theoretical and clinical significance for the study of visual neglect.
PS-100 and NF 70-200 Double Immunolabeling for Human Digital Skin Meissner Corpuscle 3D Imaging
Authors: Guinard Didier , Usson Yves , Guillermet Christiane , Saxod Raymond
Published: April 2011

For detailed study of complex structures such as corpuscular mechanoreceptors, confocal microscopy can be used with multiple immunolabeling that identifies specifically different subcomponents. In addition, anatomic interpretation is enhanced by three-dimensional reconstruction. Confocal laser micrographs, reconstructed from serial images 1 μm thick of human skin Meissner corpuscles simultaneously immunostained for neurofilaments (NF 70-200) and protein S-100 (PS-100), clearly reveal the complex 3D relationship between Schwann-related lamellar cells immunoreactive for PS-100 and the nerve fibers marked by NF 70-200. The nerve fiber, after branching into the corpuscle, divides into several ramifications, presenting discoidal expansions and flattened fringed sections. The mean nerve diameter was 4 μm ± 1 (2-5 μm) and the mean size of the discoidal expansions was 15 μm ± 1 (7-30 μm). Corpuscle size varied from 30-140 ± 1 μm in length and from 20-60 ± 1 μm in diameter. This study confirms the presence of neural discoidal areas in Meissner's corpuscles, which are probably involved to some extent with the transduction process. Despite the accuracy of immunolabeling and imaging, an extracorpuscular neural network was never observed in the vicinity of corpuscles, thus giving doubt as to their existence.
Unaltered Distribution of Laminins, Fibronectin, and Tenascin in Celiac Intestinal Mucosa
Authors: Korhonen Matti , Ormio Mia , Burgeson Robert E. , Virtanen Ismo , Savilahti Erkki
Published: April 2011

Extensive remodeling of the extracellular matrix (ECM) occurs in inflammatory tissues. The celiac lesion in the small intestine is characterized by inflammation accompanied by profound morphological alterations. We used immunohistochemistry to determine the distribution of laminin, fibronectin, and tenascin isoforms in small intestinal biopsies of untreated patients with celiac disease. In normal mucosa, the distribution of laminin isoforms defines three epithelial basement membrane (BM) zones. We found that the organization of these zones was maintained in the celiac mucosa. Thus, components of laminin-5 (α3 and β3) were found in the surface epithelial BM, laminin α2 chain was found selectively at crypt bottoms, and laminin α5 chain was the sole α-type chain in middle crypt BMs. Likewise, the distribution of fibronectin and tenascin resembled that of the normal gut. The organization of pericryptal fibroblasts and lamina propria smooth muscle strands, as defined by immunostaining for α-smooth muscle actin, also remained unchanged in the celiac mucosa. Unexpectedly, major ECM changes were not detected in the celiac lesion.
Isolation and Expression of PASK, a Serine/Threonine Kinase, During Rat Embryonic Development, with Special Emphasis on the Pancreas
Authors: Miao Ningning , Fung Brenda , Sanchez Ricardo , Lydon John , Barker Doug , Pang Kevin
Published: April 2011

We report the isolation and characterization of a serine/threonine kinase expressed during rat pancreas development. This kinase was cloned as part of a general screen using degenerate oligonucleotides to map expression of kinases and receptors during the course of pancreatic development. Sequence analysis showed it to be a member of the ste20-like serine/threonine kinase family. Northern blotting analysis against both fetal and adult tissues showed two transcripts, one of 2 kb and the other of 4 kb. The ratio of transcript expression varied with the tissue. In situ hybridization analysis showed that this gene is expressed in the early gut and pancreatic epithelium. By embryonic Day 15, the transcript is localized to cells that will eventually become exocrine in nature. In situ hybridization analysis also demonstrated high levels of expression in the choroid plexus, the developing myocardium, kidney, CNS, dorsal root ganglia, and testes. In addition, a search of the EST database revealed a related human kinase not previously described.
Immunolocalization of Anion Exchanger AE2, Na+/H+ Exchangers NHE1 and NHE4, and Vacuolar Type H+-ATPase in Rat Pancreas
Authors: Roussa Eleni , Alper Seth L. , Thévenod Frank
Published: April 2011

We have studied the expression and localization of several H+ and HCO3– transporters, whose presence in the rat pancreas is still unclear. The Cl–/HCO3– exchanger AE2, the Na+/H+ exchangers NHE1 and NHE4, and the 31-kD and 70-kD vacuolar H+-ATPase (V-ATPase) subunits were detected by immunoblotting and immunocytochemical techniques. Immunoblotting of plasma membranes with transporter-specific antibodies revealed protein bands at ≈160 kD for AE2, at ≈90 kD and ≈103 kD for NHE1 and NHE4, respectively, and at 31 kD and 70 kD for V-ATPase. NHE1 and NHE4 were further identified by amplification of isoform-specific cDNA using RT-PCR. Immunohistochemistry revealed a basolateral location of AE2, NHE1, and NHE4 in acinar cells. In ducts, NHE1 and NHE4 were basolaterally located but no AE2 expression was detected. V-ATPase was detected in zymogen granules (ZGs) by immunogold labeling, and basolaterally in duct cells by immunohistochemistry. The data indicate that NHE1 and NHE4 are co-expressed in rat pancreatic acini and ducts. Basolateral acinar AE2 could contribute to Cl– uptake and/or pH regulation. V-ATPase may be involved in ZG fusion/exocytosis and ductal HCO3– secretion. The molecular identity of the ductal Cl–/HCO3– exchanger remains unclear.
Protease XXIV Increases Detection of Mucin Gene Expression during In Situ Hybridization in Archival Tissue
Authors: Rheinhardt Jeanette M. , Finkbeiner Walter E.
Published: April 2011

Digoxigenin-labeled riboprobes of six groups of human mucins were evaluated for sensitivity in archival tissue, using protease XXIV or proteinase K during in situ hybridization.
Quantitative Cytochemical Analysis of Glucose-6-phosphate Dehydrogenase Activity in Living Isolated Hepatocytes of European Flounder for Rapid Analysis of Xenobiotic Effects
Authors: Winzer Katja , Van Noorden Cornelis J.F. , Köhler Angela
Published: April 2011

There is a great need for rapid but reliable assays to determine quantitatively effects of xenobiotics on biological systems in environmental research. Hepatocytes of European flounder are sensitive to low-dose toxic stress. Glucose-6-phosphate dehydrogenase (G6PDH) is the major source of NADPH in cells and is therefore of major importance for NADPH-dependent xenobiotic biotransformation and defense against toxic injury. These facts prompted us to develop a sensitive cytochemical method to detect G6PDH activity in living isolated flounder hepatocytes using the tetrazolium salt method. The intact plasma membrane did not appear to be a barrier for substrate, co-enzyme, and dye molecules because the intracellular enzyme reaction started immediately when incubation medium was added and could be monitored in real time per individual cell using image analysis. The reaction was effectively stopped for end point measurements by using 4% formaldehyde in 0.1 M phosphate buffer (pH 5.3). The final reaction product, formazan, was stable in hepatocytes for at least 12 days at 4C. This is the first time that a chromogenic histochemical assay is applied to living cells. This approach provides an easy tool for large-scale screening of xenobiotic metabolism and cellular stress defense. (J Histochem Cytochem 49:1025–1032, 2001)
Monitoring Signal Transduction in Cancer
Authors: Hsieh H.-Ben , Lersch Robert A. , Callahan Daniel E. , Hayward Simon , Wong Mariwil , Clark Orlo H. , Weier Heinz-Ulrich G.
Published: April 2011

This study targeted the development of a novel microarray tool to allow rapid determination of the expression levels of 58 different tyrosine kinase (tk) genes in small tumor samples. The goals were to define a reference probe for multi-sample comparison and to investigate the variability and reproducibility of the image acquisition and RT-PCR procedures. The small number of tk genes on our arrays enabled us to define a reference probe by artificially mixing all genes on the arrays. Such a probe provided contrast reference for comparative hybridization of control and sample DNA and enabled cross-comparison of more than two samples against one another. Comparison of signals generated from multiple scanning eliminated the concern of photo bleaching and scanner intrinsic noise. Tests performed with breast, thyroid, and prostate cancer samples yielded distinctive patterns and suggest the feasibility of our approach. Repeated experiments indicated reproducibility of such arrays. Up- or downregulated genes identified by this rapid screening are now being investigated with techniques such as in situ hybridization. (J Histochem Cytochem 49:1057–1058, 2001)
Detection of the Mr 110,000 Lung Resistance-related Protein LRP/MVP with Monoclonal Antibodies
Authors: Schroeijers Anouk B. , Scheffer George L. , Reurs Anneke W. , Pijnenborg Adriana C.L.M. , Abbondanza Ciro , Wiemer Erik A.C. , Scheper Rik J.
Published: April 2011

The Mr 110,000 lung resistance-related protein (LRP), also termed the major vault protein (MVP), constitutes >70% of subcellular ribonucleoprotein particles called vaults. Overexpression of LRP/MVP and vaults has been linked directly to MDR in cancer cells. Clinically, LRP/MVP expression can be of value to predict response to chemotherapy and prognosis. Monoclonal antibodies (MAbs) against LRP/MVP have played a critical role in determining the relevance of this protein in clinical drug resistance. We compared the applicability of the previously described MAbs LRP-56, LMR-5, LRP, 1027, 1032, and newly isolated MAbs MVP-9, MVP-16, MVP-18, and MVP-37 for the immunodetection of LRP/MVP by immunoblotting analysis and by immunocyto- and histochemistry. The availability of a broader panel of reagents for the specific and sensitive immunodetection of LRP/MVP should greatly facilitate biological and clinical studies of vault-related MDR.
CP27 Localization in the Dental Lamina Basement Membrane and in the Stellate Reticulum of Developing Teeth
Authors: Diekwisch Thomas G.H. , Luan Xianghong , McIntosh James E.
Published: April 2011

cp27 is a novel gene involved in early vertebrate development that features a distinct protein localization pattern in developing tooth organs. During initial tooth development, CP27 was detected at the epithelial-mesenchymal interface of dental lamina stage tooth organs. At later stages of tooth development, CP27 was localized in the stellate reticulum, the oral mucosa mesenchyme, and alveolar bone. The significant changes in the highly restricted distribution pattern suggest that CP27 might be involved at several different levels during tooth development.
Keratan Sulfate Epitopes Exhibit a Conserved Distribution During Joint Development That Remains Undisclosed on the Basis of Glycosaminoglycan Charge Density
Authors: Kavanagh Emma , Osborne Anne C. , Ashhurst Doreen E. , Pitsillides Andrew A.
Published: April 2011

Changes in glycosaminoglycan (GAG) content and distribution are vital for joint development. However, their precise character has not been established. We have used immunohistochemistry (IHC) and “critical electrolyte” Alcian blue staining to assess such changes in developing chick and rabbit joints. IHC showed chondroitin sulfate labeling in chick epiphyseal cartilage but not in interzones. In contrast, prominent labeling for keratan sulfate (KS) was restricted to chick cartilage–interzone interfaces. In rabbit knees, KS labeling was also prominent at presumptive cavity borders, but weak in interzone and cartilage. Selective pre-digestion produced appropriate loss of label and undersulfated KS was undetectable. Quantification of Alcian blue staining by scanning and integrating microdensitometry showed prominent hyaluronan-like (HA-like) interzone staining, with chondroitin sulfate and weaker KS staining restricted to epiphyseal cartilage. Hyaluronidase decreased HA-like staining in the interzone. Surprisingly, keratanases also reduced HA-like but not sulfated GAG (sGAG-like) staining in the interzone. Chondroitinase ABC had little effect on HA-like staining but decreased sGAG staining in all regions. Rabbit joints also showed HA-like but not KS staining in the interzone and strong chondroitin sulfate-like staining in epiphyseal cartilage. Our findings show restricted KS distribution in the region close to the presumptive joint cavity of developing chick and rabbit joints. Alcian blue staining does not detect this moiety. Therefore, it appears that although histochemistry allows relatively insensitive quantitative assessment of GAGs, IHC increases these detection limits. This is particularly evident for KS, which exhibits immunolabeling patterns in joints from different species that is consistent with a conserved functional role in chondrogenesis.
A Quantitative Method to Determine the Orientation of Collagen Fibers in the Dermis
Authors: Noorlander Maril L. , Melis Paris , Jonker Ard , Noorden Cornelis J.F. Van
Published: April 2011

We have developed a quantitative microscopic method to determine changes in the orientation of collagen fibers in the dermis resulting from mechanical stress. The method is based on the use of picrosirius red-stained cryostat sections of piglet skin in which collagen fibers reflect light strongly when epipolarization microscopy is used. Digital images of sections were converted into binary images that were analyzed quantitatively on the basis of the length of the collagen fibers in the plane of the section as a measure for the orientation of the fibers. The length of the fibers was expressed in pixels and the mean length of the 10 longest fibers in the image was taken as the parameter for the orientation of the fibers. To test the procedure in an experimental setting, we used skin after 0 and 30 min of skin stretching. The orientation of the fibers in sections of control skin differed significantly from the orientation of fibers in sections of skin that was stretched mechanically for 30 min [76 ± 15 ( n = 5) vs 132 ± 36 ( n = 5)]. The method described here is a relatively simple way to determine (changes in) the orientation of individual collagen fibers in connective tissue and can also be applied for analysis of the orientation of any other structural element in tissues so long as a representative binary image can be created.
An Improved Protocol of Biotinylated Tyramine-based Immunohistochemistry Minimizing Nonspecific Background Staining
Authors: Kim Seok Hyung , Shin Young Kee , Lee Kyung Mee , Lee Jung Sun , Yun Ji Hye , Lee Sun Myung
Published: April 2011

An immunohistochemical method using biotinyl tyramine was recently introduced to amplify weak staining signals. Despite its high sensitivity, however, tyramine-based immunostaining has been limited by its increased background staining. In this study, to develop an improved protocol of biotinyl tyramine-based immunohistochemistry minimizing the background staining, we determined which staining steps lead to the nonspecific reaction and the most appropriate blocking agents for background-provoking steps. Trypton casein peptone and distilled water with Tween-20 were shown to be most effective as a blocking agent and a rinsing solution, respectively. In conclusion, we developed an optimized protocol for biotinyl tyramine-based immunohistochemistry with minimal background staining.
Selective Binding of Lectins to Embryonic Chicken Vasculature
Authors: Jilani Shahla M. , Murphy Thomas J. , Thai Shelley N.M. , Eichmann Anne , Alva Jackelyn A. , Iruela-Arispe M. Luisa
Published: April 2011

Chicken embryos are an excellent model system for studies related to vascular morphogenesis. Development in ovo allows manipulations otherwise difficult in mammals, and the use of chicken-quail chimeras offers an additional advantage to this experimental system. Furthermore, the chicken chorioallantoic membrane has been extensively used for in vivo assays of angiogenesis. Surprisingly, few markers are available for a comprehensive visualization of the vasculature. Here we report the use of lectins for identification of embryonic chicken blood vessels. Nine lectins were evaluated using intravascular perfusion and directly on sections. Our results indicate that Lens culinaris agglutinin, concanavalin A, and wheat germ agglutinin can be used effectively for visualization of vessels of early chicken embryos (E2.5-E4). At later developmental stages, Lens culinaris agglutinin is a better choice because it displays equal affinity for the endothelia of arteries, veins, and capillaries. The findings presented here expand our understanding of lectin specificity in the endothelium of avian species and provide information as to the use of these reagents to obtain comprehensive labeling of the embryonic and chorioallantoic membrane vasculature.
Upregulation of Cardiac Insulin-like Growth Factor-I Receptor by ACE Inhibition After Myocardial Infarction: Potential Role in Remodeling
Authors: Dean Rachael G. , Bach Leon A. , Burrell Louise M.
Published: April 2011

This study evaluated the effects of angiotensin-converting enzyme (ACE) inhibition after myocardial infarction (MI) on cardiac remodeling and gene expression and localization of components (ligands, receptors, and binding proteins) of the cardiac insulinlike growth factor (IGF) system. After ligation of the coronary artery, rats were randomized to vehicle or ACE inhibitor (Captopril, 50 mg/kg/day) for 4 weeks. Blood pressure, cardiac remodeling, and components of the IGF system were localized in the heart using in situ hybridization (ISH) and immunohistochemistry (IHC). The average infarct size was 42%. There were regional differences in the expression of the IGF system after MI, with increased IGF-I mRNA abundance in the border (24-fold) and infarct (12-fold) and increased IGF-binding protein (IGFBP)-3 mRNA in all areas of the failing left ventricle (threefold). Captopril reduced blood pressure, attenuated cardiac remodeling, and caused a threefold increase in IGF-I receptor mRNA and protein in infarct, border zone, and viable myocardium ( p<0.01). Captopril had no effect on IGF-I mRNA but further increased IGFBP-3 mRNA and protein in the border zone, ( p<0.05). The changes in the cardiac IGF system following MI are highly localized, persist for at least 4 weeks, and can be modulated by ACE inhibition. These data suggest that the benefits of ACE inhibitors in attenuation of cardiac remodeling may be mediated in part through increased expression of the IGF-I receptor sensitizing the myocardium to the positive effects of endogenous IGF-I.