Mouse Sperm Rosette: Assembling During Epididymal Transit,in vitro Disassemble,and Oligosaccharide Participation in the Linkage Material

In many mammals, sperm associations had been observed, but not in the mouse. In this work, mouse sperm rosettes are morphologically described inside the epididymis and during its dissolution in a culture medium. Also characterized are the saccharides present in the linking material. Sperm association and other epididymal actions are supported by sperm during epididymal transit and are verified at the caudal region, suggesting a relation between epididymal transit and sperm maturation. In drops of epididymal content obtained from distal (cauda), but not from proximal (caput and corpus) regions; dissolved in culture medium, rosettes appear to be 10 to 15 motile sperm joined by their heads. After 3 min, sperm progressively detach, disassembling the rosette. These structures are studied by several techniques, including optic, electronic (scanning electron microscopy and transmission electron microscopy), and video microscopy. At the ultrastructural level, a dense network of electron‐dense material was observed between sperm heads, joining them. Based on previous works in rat, several lectins were used to characterize the type of saccharides present in this linking material. To avoid the contact between sperm and epididymal fluid from distal region—that probably exerts an influence on sperm association—a ligature was placed between caput and corpus. This epididymal content isolated from caput did not display any rosettes after 28 days. Anat Rec, 2007. © 2007 Wiley‐Liss, Inc.     

Integrin expression and osteopontin regulation in human fetal osteoblastic cells mediated by substratum surface characteristics

Integrin-mediated adhesion of anchorage-dependent cells to scaffolds is a critical component of tissue engineering. We investigated integrin expression by the human fetal osteoblastic cell line, hFOB 1.19 (hFOB), as a function of substratum surface wettability. The influence of surface wettability on bone cell phenotype was also examined. Plasma-treated quartz (PTQ) and glass (PTG) (hydrophilic, contact angles of 0 degrees), octadecyltrichlorosilane-treated quartz (STQ) and glass (STG) (hydrophobic, contact angles above about 100 degrees), and tissue culture polystyrene were used for cell culture. hFOB cells cultured on hydrophilic substrata displayed well-developed actin stress fibers relative to cells on hydrophobic substrata. Western blot analysis revealed that hFOB cells cultured on hydrophobic substrata (STQ or STG) express lower levels of alphav and beta3 integrin subunits than do cells on hydrophilic substrata (PTQ or PTG). This effect was more pronounced in cells on STQ than on STG. These variations in integrin expression were lessened by extended culture time. Double- labeled integrin/actin immunofluorescence confirmed Western blot results, that is, cells cultured on PTQ displayed distinct, large plaques of alphav and beta3 subunits and integrin alphavbeta3, as well as their colocalization with actin stress fiber ends, whereas cells on STQ did not display integrin plaques after 24 h and displayed only minimal plaque formation after 3 days. Vinculin, a focal adhesion protein that mediates binding between the integrin and actin cytoskeleton, appeared in Western blots to mimic the variations of alphav and beta3 expression with respect to surface wettability. Interestingly, real-time RT-PCR analysis showed that hFOB cultured on hydrophobic substrata, which have downregulated alphav and beta3 integrin subunits, displayed greater steady state mRNA levels of osteopontin, an extracellular matrix (ECM) protein containing the Arg-Gly-Asp (RGD) integrin recognition sequence, than did cells cultured on hydrophilic substrata. Our results imply that substratum surface wettability regulates integrin-mediated bone cell adhesion and further influences the expression of bone cell-ECM complexes.     

Oxygen Tension Imaging in the Mouse Retina

A newly developed microscope-based imaging system was used to measure the oxygen tension (PO₂) inside the retinal and choroidal vessels of mice and to generate in vivo maps of retinal PO₂. These maps were generated from the phosphorescence lifetimes of an injected palladium–porphyrin compound using a frequency-domain measurement. The system was fully calibrated and used to produce retinal PO₂ maps at different inspiratory oxygen fractions. PO₂ rose accordingly and predictably as inspiratory O₂ was stepped from hypoxic to hyperoxic conditions. Important experimental and acquisition parameters necessary for applying phosphorescence lifetime imaging to the mouse eye were investigated, including camera exposure and intensifier gain settings. Because of a need to limit light exposure to the retina, PO₂ map quality as measured by the coefficient of determination was investigated as a function of signal-to-noise and accumulated excitation energy deposition. With the development of this technology for use in mice, the potential for investigating the oxygen dynamics in genetically engineered mouse models of retinal disease, including diabetic retinopathy, glaucoma, and age-related macular degeneration, is advanced. © 2003 Biomedical Engineering Society. PAC2003: 4266Ew, 8763Lk, 8719Dd     

Physical Similarity and Twin Resemblance for Eating Attitudes and Behaviors: A Test of the Equal Environments Assumption

The Equal Environments Assumption (EEA) in twin studies of eating pathology was investigated by examining the hypothesis that twin resemblance for eating attitudes and behaviors is affected by their degree of physical similarity. Eating attitudes and behaviors were assessed in 338 female adolescent twin pairs with a revised version of the Eating Disorder Inventory (EDI). General physical similarity as well as body size/shape similarity were assessed using ratings of color photographs, ratings of body shape, and body mass index. All physical similarity assessments were conducted blind to twin zygosity. Significant associations between physical similarity and twin similarity for eating attitudes and behaviors were not found. Mean EDI within-twin pair absolute difference scores did not differ significantly among more versus less physically-similar groups. Additionally, correlation and regression analyses failed to find a significant association between EDI absolute difference scores and physical similarity indices. The current findings provide support for the EEA in twin studies of eating attitudes and behaviors.     

Lifestyle Risk Score: handling missingness of individual lifestyle components in meta-analysis of gene-by-lifestyle interactions

Recent studies consider lifestyle risk score (LRS), an aggregation of multiple lifestyle exposures, in identifying association of gene-lifestyle interaction with disease traits. However, not all cohorts have data on all lifestyle factors, leading to increased heterogeneity in the environmental exposure in collaborative meta-analyses. We compared and evaluated four approaches (Naïve, Safe, Complete and Moderator Approaches) to handle the missingness in LRS-stratified meta-analyses under various scenarios. Compared to “benchmark” results with all lifestyle factors available for all cohorts, the Complete Approach, which included only cohorts with all lifestyle components, was underpowered due to lower sample size, and the Naïve Approach, which utilized all available data and ignored the missingness, was slightly inflated. The Safe Approach, which used all data in LRS-exposed group and only included cohorts with all lifestyle factors available in the LRS-unexposed group, and the Moderator Approach, which handled missingness via moderator meta-regression, were both slightly conservative and yielded almost identical p values. We also evaluated the performance of the Safe Approach under different scenarios. We observed that the larger the proportion of cohorts without missingness included, the more accurate the results compared to “benchmark” results. In conclusion, we generally recommend the Safe Approach, a straightforward and non-inflated approach, to handle heterogeneity among cohorts in the LRS based genome-wide interaction meta-analyses.Subject terms: Genetics, Risk factors     

Microhardness of superficial and deep sound human dentin

Our purpose in this study was to determine the microhardness of superficial and deep dentin by means of two indentation methods (Knoop and Vickers) under two different applied loads. Twelve dentin discs approximately 2-mm thick were obtained from both superficial and deep dentin by transversally sectioning the crowns of sound, extracted human third molars with a diamond blade under water irrigation. Dentin surfaces were sequentially polished, and indentations (n = 20 per surface) were performed with either Vickers indentor at loads of 300 and 500 g, respectively, or Knoop indentor at loads of 50 and 100 g, respectively. Average Vickers hardness number (VHN) and Knoop hardness number (KHN) were calculated and treated with two-way analysis of variance (ANOVA) and Student's t test. Microhardness of dentin was not influenced by the different loads applied for both indentation methods. Knoop hardness was significantly higher for superficial than for deep dentin (p < 0.05). Conversely, Vickers hardness was not significantly different for both substrates (p > 0.05). Differences in dentin hardness as a function of depth exist, but they might not be relevant, and no alteration of the distribution of stresses along the adhesive interface is expected.     

Comparative analysis of immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay using virus-like particles or virus-infected mouse brain antigens to detect IgM antibody in sera from patients with evident flaviviral infections

The use of immunoglobulin M (IgM) antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA) serves as a valuable tool for the diagnosis of acute flaviviral infections, since IgM antibody titers are detectable early, peak at about 2 weeks postinfection, and subsequently decline to lower levels over the next few months. Traditionally, virus-infected tissue culture or suckling mouse brain (SMB) has been the source of viral antigens used in the assay. In an effort to provide a reliable source of standardized viral antigens for serodiagnosis of the medically important flaviviruses, we have developed a eukaryotic plasmid vector to express the premembrane/membrane and envelope proteins which self-assemble into noninfectious virus-like particles (VLPs). In addition to the plasmids for Japanese encephalitis virus, West Nile virus (WNV), St. Louis encephalitis virus (SLEV), and dengue virus type 2 (DENV-2) reported earlier, we recently constructed the DENV-1, -3, and -4 VLP expression plasmids. Three blind-coded human serum panels were assembled from patients having recent DENV, SLEV, and WNV infections to assess the sensitivity and specificity of the MAC-ELISA using VLPs or SMB antigens. In addition, serum specimens from patients infected with either Powassan virus or La Crosse encephalitis virus were used to evaluate the cross-reactivity of seven mosquito-borne viral antigens. The results of the present studies showed higher sensitivity when using SLEV and WNV VLPs and higher specificity when using SLEV, WNV, and the mixture of DENV-1 to -4 VLPs in the MAC-ELISA than when using corresponding SMB antigens. Receiver operating characteristic (ROC) curve analysis, a plot of the sensitivity versus false positive rate (100 - specificity), was applied to discriminate the accuracy of tests comparing the use of VLPs and SMB antigen. The measurement of assay performance by the ROC analysis indicated that there were statistically significant differences in assay performance between DENV and WNV VLPs and the respective SMB antigens. Additionally, VLPs had a lower cutoff positive/negative ratio than corresponding SMB antigens when employed for the confirmation of current infections. The VLPs also performed better than SMB antigens in the MAC-ELISA, as indicated by a higher positive prediction value and positive likelihood ratio test. Cell lines continuously secreting these VLPs are therefore a significantly improved source of serodiagnostic antigens compared to the traditional sources of virus-infected tissue culture or suckling mouse brain.     

The Effect of Accuracy of Perceptions of Dietary-Fat Intake on Perceived Risk and Intentions to Change

Consumption of excess fat increases risk for many health problems and diseases. In the present study, 188 undergraduate students were studied to understand self-perceptions of dietary-fat intake and the impact of those perceptions. Findings indicated that the majority of participants had inaccurate perceptions about the amount of fat in their diets. Further, compared to people who overestimated dietary-fat intake, people who underestimated fat intake had lower perceived risk of cancer, had lower intentions to change, and demonstrated less knowledge about the dietary-fat content of many foods. Findings suggest that this unrealistic underestimation of fat intake is a cognitive barrier to dietary change and people who underestimate dietary fat intake may require more intensive intervention to change their diets.     

Diagnostic performance of arginase activity in colorectal cancer

Arginase activity was measured in serum and biopsy from healthy individuals and colorectal cancer patients. Arginase activity in tumor samples (87±7.7 U/g tissue) was significantly higher than in controls (40.7±3.3 U/g tissue). However, serum arginase activity did not show any significant change in both groups. Finally, the micromethod used to quantify arginase activity in this study is superior to other methods because it has increased sensitivity, requires less sample, and is less time-consuming. Arginase differences are significant, according to the t-test (P<0.05). Received: 14 September 2001 / Accepted: 6 February 2002