Magnetic Resonance Imaging Assessment of Subchondral Bone and Soft Tissues in Knee Osteoarthritis

Knee osteoarthritis (OA) has to be considered a whole joint disease. Magnetic resonance imaging (MRI) allows superior assessment of all joint tissues that may be involved in OA, such as the subchondral bone, synovium, ligaments, and periarticular soft tissues. Reliable MRI-based scoring systems are available to assess and quantify these structures and associated pathology. Cross-sectional and longitudinal evaluation has enabled practitioners to understand their relevance in explaining pain and structural progression.

Magnetic Resonance Imaging in Knee Osteoarthritis Research: Semiquantitative and Compositional Assessment

Semiquantitative assessment of the knee by expert magnetic resonance imaging readers is a powerful research tool for understanding the natural history of osteoarthritis (OA). Several reliable semiquantitative scoring systems have been applied to large observational cross-sectional and longitudinal epidemiologic studies and interventional clinical trials. Such evaluations have enabled understanding of the relevance of disease in structures within the knee joint to explain pain and progression of OA. Compositional imaging of cartilage has added to our ability to detect early degeneration before morphologic changes are present, which may help to prevent the permanent morphologic changes commonly seen in knee OA.

Number of expressed cancer/testis antigens identifies focal adhesion pathway genes as possible targets for multiple myeloma therapy

Considering that the importance of cancer/testis (CT) antigens in multiple myeloma (MM) biology is still under investigation, the present study aimed to: (1) identify genes differentially expressed in MM using microarray analysis of plasma cell samples, separated according to the number of expressed CTs; (2) examine possible pathways related to MM pathogenesis; (3) validate the expression of candidate genes by quantitative real-time PCR (RQ-PCR). Three samples predominantly positive (>6 expressed), including the U266 cell line, and three samples predominantly negative (0 or 1 expressed CT for the 13 analyzed CT antigens), were submitted for microarray analysis. Validation by RQ-PCR from 24 MM samples showed that the ITGAS gene was downregulated in predominantly positive (>6 expressed CTs, p = 0.0030) and in tumor versus normal plasma cells (p = 0.0182). The RhoD gene was overexpressed in tumor plasma cells when compared to normal plasma cells (p = 0.0339). Results of the microarray analysis corroborate the hypothesis that MM could be separated into predominantly positive and predominantly negative expression. The differential expression of ITGA5 and RhoD suggests disruption of the focal adhesion pathway in MM and offers a new target field to be explored in this disease.

Tissue diffusion and retention of metalloproteinases in ascending aortic aneurysms and dissections

Histopathological alterations in human aneurysms and dissections of the thoracic ascending aorta include areas of mucoid degeneration within the medial layer, colocalized with areas of cell disappearance and disruption of extracellular matrix elastic and collagen fibers. We studied the presence of matrix metalloproteinases in relation to their capacity to diffuse through the tissue or to be retained in areas of mucoid degeneration in aneurysms and dissections of the ascending aorta. Ascending aortas from 9 controls, 33 patients with aneurysms, and 14 with acute dissections, all collected at surgery, were analyzed. The morphological aspect was similar whatever the etiology or phenotypic expression of the pathological aortas, involving areas of extracellular matrix breakdown and cell rarefaction associated with mucoid degeneration. Release of proMMP-2, constitutively expressed by smooth muscle cells, was not different between controls and aneurysmal aortas, whereas the aneurysmal aortas released more of the active form. Release of pro and active MMP-9 was also similar between controls and aneurysmal aortas. Immunohistochemical staining of MMP-2 and MMP-9 was weak in both control and pathological aortas. In contrast, released MMP-7 (matrilysin) and MMP-3 (stromelysin-1) could not be detected in conditioned media but were present in tissue extracts with no detectable quantitative difference between controls and pathological aortas. Immunohistochemical staining of MMP-7 and MMP-3 revealed their retention in areas of mucoid degeneration, and semiquantitative evaluation of immunostaining showed more MMP-7 in pathological aortas than in controls. In conclusion, areas of mucoid degeneration, the hallmark of aneurysms, and dissections of thoracic ascending aortas, whatever their etiology, are not inert and can retain specific proteases. (c) 2009 Elsevier Inc. All rights reserved.

Hyperkalemia: A quick reference

This article serves as a quick reference for hyperkalemia. Guidelines for analysis and causes, signs, and a stepwise approach are presented.

Hypokalemia: A quick reference

This article serves as a quick reference for hypokalemia. Guidelines for analysis and causes, signs, and a stepwise approach are presented.

Lsa63, a newly identified surface protein of Leptospira interrogans binds laminin and collagen IV

Leptospira interrogans is the etiological agent of leptospirosis, a zoonotic disease that affects populations worldwide. We have identified in proteomic studies a protein that is encoded by the gene LIC10314 and expressed in virulent strain of L. interrogans serovar Pomona. This protein was predicted to be surface exposed by PSORT program and contains a p83/100 domain identified by BLAST analysis that is conserved in protein antigens of several strains of Borrelia and Treponema spp. The proteins containing this domain have been claimed antigen candidates for serodiagnosis of Lyme borreliosis. Thus, we have cloned the LIC10314 and expressed the protein in Escherichia coli BL21-SI strain by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and characterized by circular dichroism spectroscopy. This protein is conserved among several species of pathogenic Leptospira and absent in the saprophytic strain L. biflexa. We confirm by liquid-phase immunofluorescence assays with living organisms that this protein is most likely a new surface leptospiral protein. The ability of the protein to mediate attachment to ECM components was evaluated by binding assays. The leptospiral protein encoded by LIC10314, named Lsa63 (Leptospiral surface adhesin of 63 kDa), binds strongly to laminin and collagen IV in a dose-dependent and saturable fashion. In addition, Lsa63 is probably expressed during infection since it was recognized by antibodies of serum samples of confirmed-leptospirosis patients in convalescent phase of the disease. Altogether, the data suggests that this novel identified surface protein may be involved in leptospiral pathogenesis. (C) 2009 The British Infection Society. Published by Elsevier Ltd. All rights reserved.

Lp95, a novel leptospiral protein that binds extracellular matrix components and activates e-selectin on endothelial cells

Objectives: The study of a predicted outer membrane leptospiral protein encoded by the gene LIC12690 in mediating the adhesion process. Methods: The gene was cloned and expressed in Escherichia coli BL21 (SI) strain by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and used to assess its ability to activate human umbilical vein endothelial cells (HUVECs). Results: The recombinant leptospiral protein of 95 kDa, named Lp95, activated E-selectin in a dose-dependent fashion but not the intercellular adhesion molecule 1 (ICAM-1). In addition, we show that pathogenic and non-pathogenic Leptospira are both capable to stimulate endothelium E-selectin and ICAM-1, but the pathogenic L. interrogans serovar Copenhageni strain promotes a statistically significant higher activation than the non-pathogenic L. biflexa serovar Patoc (P < 0.01). The Lp95 was identified in vivo in the renal tubules of animal during experimental infection with L. interrogans. The whole Lp95 as well as its fragments, the C-terminal containing the domain of unknown function (DUF), the N-terminal and the central overlap regions bind laminin and fibronectin ECM molecules, being the binding stronger with the DUF containing fragment. Conclusion: This is the first leptospiral protein capable to mediate the adhesion to ECM components and the activation of HUVECS, thus suggesting its participation in the pathogenesis of Leptospira. (C) 2009 The British Infection Society. Published by Elsevier Ltd. All rights reserved.

Exogenous DNA uptake by bovine spermatozoa does not induce DNA fragmentation

Sperm-mediated gene transfer (SMGT) is a fast and low-cost method used to produce transgenic animals. The objective of this study was to evaluate the effects of the concentration of exogenous DNA and the duration of incubation on DNA uptake by bovine spermatozoa and subsequently the integrity of sperm DNA and sperm apoptosis. Spermatozoa (5 X 10(6) cells/mL) were incubated with 100, 300, or 500 ng of exogenous DNA (pEYFP-Nuc plasmid) for 60 or 120 min at 39 degrees C. The amount of exogenous DNA associated with spermatozoa was quantified by real-time PCR, and the percentages of DNA fragmentation in spermatozoa were evaluated using SCSA and a TUNEL assay, coupled with flow cytometry. Uptake of exogenous DNA increased significantly as incubation increased from 60 to 120 min (0.0091 and 0.028 ng, respectively), but only when the highest exogenous DNA concentration (500 ng) was used (P < 0.05). Based on SCSA and TUNEL assays, there was no effect of exogenous DNA uptake or incubation period on sperm DNA integrity. In conclusion, exogenous DNA uptake by bovine spermatozoa was increased with the highest exogenous DNA concentration and longest incubation period, but fragmentation of endogenous DNA was apparently not induced. (C) 2010 Elsevier Inc. All rights reserved.

Sequence variation of the alpha-lactalbumin gene in Holstein and Nellore cows

The alpha-lactalbumin is a subunit of lactose-synthase, an enzyme responsible for lactose production, a disaccharide that influences milk production. Sequence variations of bovine alpha-lactalbumin have been associated with differences in milk yield. This study aimed to analyze allelic frequency differences at position-1689 (g. AG) and+15 (g. AG) of the alpha-lactalbumin gene in Holstein (Bos taurus) and Nellore (Bos indicus) cows. Blood samples were analyzed from 34 Holstein, 104 Nellore, and 99 Dairy Nellore cows using PCR-RFLP. The different RFLP patterns were sequenced and a novel sequence variation on nucleotide-46 was identified. An adenine at this position was designated as the A allele and a guanine was designated B allele. The frequencies of alleles A-1689, A-46, and A+15 differed between Holstein and both Nellore breeds. The results show that differences in alpha-lactalbumin allelic variants in the 5`-flanking and the 5`-UTR region might be associated with differences in milk production between Holstein cows and cows from Nellore breeds. However, the lack of difference between Nellore and Dairy Nellore suggests that other sequence variantions that regulate milk production might be responsible for the selection of Dairy Nellore cows with superior milk production.

Vitrification with Glutamine Improves Maturation Rate of Vitrified/Warmed Immature Bovine Oocytes

Contents The current study examined the protective effects of l-glutamine and cytochalasin B during vitrification of immature bovine oocytes. Oocyte vitrification solution (PBS supplemented with 10% FCS, 25% EG, 25% DMSO and 0.5 m trehalose) was the vitrification control. Treatments were the addition of 7 mu g/ml cytochalasin B, 80 mm glutamine or both cytochalasin and glutaminine for 30 s. After warming, oocytes were matured in vitro for 24 h, fixed and stained with Hoechst (33342) for nuclear maturation evaluation. l-glutamine improved the vitrified/warmed immature bovine oocytes viability (32.8%), increasing the nuclear maturation rates compared to other treatments and the no treatment vitrified control (17.4%). There was, however, no effect of cytochalasin B on in vitro maturation (14.4%).

A Novel Approach to Intraoral Mandibular Nerve Anesthesia: Changing Reference Planes in the Gow-Gates Block Technique

Purpose: The Gow-Gates technique is said to have several advantages over traditional techniques to achieve mandibular nerve anesthesia; however, its routine use is quite limited, mainly due to complications during visual alignment of reference landmarks. The purpose of this study was to verify the validity and accuracy of a new method to reach the injection site. Material and Methods: Fifteen magnetic resonance images were captured. Distances from the ideal injection point in the condylar neck (puncture ideal) to the injection points located in the a and 0 plane intersection (Puncture Gow-Gates and puncture modified) were measured and compared. Results: Positive and significant (P <= .003) Pearson correlations between landmarks and injection points confirmed the validity of the modified technique. Paired t test showed that the segment line puncture ideal-puncture modified, 5.17 mm, was 3 times shorter (P < .001) than the segment line puncture ideal-puncture Gow-Gates, 17.91 mm. As calculated by linear regression, establishing the injection point of the modified technique depended only on the anteroposterior and lateromedial condyle positions. Conclusions: The modified technique proved to be valid and precise and has a determined and an effective injection site. (C) 2009 American Association of Oral and Maxillofacial Surgeons J Oral Maxillofac Surg 67:2609-2616, 2009

Nucleophosmin, p53, and Ki-67 expression patterns on an oral squamous cell carcinoma tissue microarray

Oral cancer is the eighth most prevalent cancer worldwide. It causes significant mortality and morbidity rates, which have motivated the search for prognostic factors to better tailor the individual management of oral squamous cell carcinoma patients. Nucleophosmin is a multifunctional protein that is involved in many cellular activities, such as, regulation of the tumor suppressor genes TP53 and p14(ARF). and is associated with proliferative and growth suppressive roles in the cell. Nucleophosmin is overexpressed in many solid tumors in human, including tumors of the colon, liver, stomach, ovary, and prostate. In this study, we analyzed the expression of nucleophosmin, Ki-67, and p53 by immunohistochemistry in oral squamous cell carcinomas. Less than 10% of nuclear staining was observed in 90.3%, 50.6%, and 65.3% of the cases for nucleophosmin, p53, and Ki-67, respectively. Expression of p53 was not significantly associated with any of the clinicopathologic parameters analyzed. Increased expression of Ki-67 was associated with the presence of lymph node metastasis (P < .0001), advanced stages of disease (P = .0030), tumors occurring in the floor of mouth (P = .0018), and moderately/well-differentiated tumors (P = .0287). Local recurrence was associated with higher expression of nucleophosmin (P = .0233), and disease-free survival rate was significantly better in patients with low expression of nucleophosmin. Multivariate analysis suggested that expression of nucleophosmin could be an independent prognostic factor for oral squamous cell carcinoma patients. (C) 2010 Elsevier Inc. All rights reserved.