NIH Imaging Research Articles

Outcome of Repeated Thrombolysis in Debrecen Thrombolysis Database

Recombinant tissue plasminogen activator (rtPA) is an efficient therapy of acute ischemic stroke. The risk of a recurrent ischemic stroke is high. This prospective single center study aimed to assess whether or not the repeated rtPA treatment is beneficial for acute stroke patients. All thrombolysed patients' data at the Department of Neurology, University of Debrecen have been recorded in the Debrecen Thrombolysis Database (DTD) since 2004. We identified 21 patients with repeated thrombolysis. Stroke severity by the NIH stroke scale score (NIHSSS) and imaging findings by the Alberta Stroke Programme Early CT Score were evaluated on admission and 1 day later. The modified Rankin Scale score at 3 months and case fatality at 1 year were evaluated. We compared the first and second thrombolyses, and we screened for bleeding and allergic reactions to determine safety. Within the 27-month median time, 18 patients were thrombolysed twice, with complete follow-up. In recurrent stroke patients, diabetes mellitus, congestive heart failure, and anticoagulation were more common. Admission cholesterol levels were decreased. After the first and second treatments, 24-hour NIHSSS were 3 (1;6) and 7 (1;10), respectively. At 3 months, good outcome was significantly higher after the first treatment than those of DTD, with no differences between the 2 attempts. There was little difference in 3-month and 1-year outcomes, regardless of laterality-ipsilateral or contralateral hemisphere-in recurrent strokes. One patient had nonsymptomatic intracranial bleeding after repeated rtPA treatment. Recurrent rtPA treatment may be safe and effective in patients who have mild or moderate residual symptoms after the index stroke.

Digital Image Analysis to Characterize the Upper Lid Marginal Peak After Levator Aponeurosis Repair

Lid contour is important for eyelid symmetry, but clinical quantification can be difficult. The marginal peak is a major determinant of upper eyelid contour, and its horizontal position, relative to the mid-pupillary line, may be used as a measure of symmetry after surgery. This study used digital image analysis to assess the peak position in patients who had undergone levator aponeurosis repair. An unselected group of patients underwent anterior-approach levator aponeurosis repair, performed under local anesthesia by a single surgeon, using 3 6/0 Vicryl sutures to reattach the levator aponeurosis to the upper tarsus. Both before and 2-3 weeks after ptosis repair, digital photographs were taken in a fixed format and the images analyzed using NIH imaging software to assess the position of the upper eyelid peak. Comparisons were made between the pre- and postoperative measurements in the operated eye and between the operated eye and its fellow eye. Twenty people (8 male; 40%) underwent unilateral levator muscle resection (9 right eyes; 45%). In eyes before ptosis surgery, the peak was significantly more medially placed (median +0.17 mm) as compared with that of fellow eyes (median +0.80 mm) (p = 0.023). There was a significant temporal shift of the median peak after surgery, from +0.17 mm to +0.92 mm (p = 0.029). Although there was a minor nasal shift of peak in the unoperated eyes after contralateral surgery (median +0.80 mm to +0.64 mm), the peak positions for the 2 eyes were not significantly different at 2-3 weeks after aponeurosis repair (p = 0.22). Digital image analysis allows an objective assessment of the upper eyelid marginal contour after ptosis repair. A nasal shift in the upper eyelid marginal peak has been identified in ptotic eyes (as compared with the contralateral side) and, after successful ptosis repair, there is a significant temporal shift to match the unoperated fellow eye.

Evaluation of a new eastern blotting technique for the analysis of ginsenoside Re in American ginseng berry pulp extracts

A new eastern blotting technique has been established for ginsenoside Re (G-Re) contained in American ginseng berry pulp extracts. G-Re in American ginseng berry pulp was extracted using 100% methanol, 100% ethanol, 50% aqueous methanol, and 50% aqueous ethanol. The combined crude extracts were applied onto a polyethersulfone membrane and developed using the methanol–water–acetic acid solvent system (45:55:1 v/v). Separated components were immunostained using anti-G-Re monoclonal antibody. G-Re was first specifically detected and then quantitatively analyzed using NIH Imaging software. We also confirmed that the most suitable solvent was 50% aqueous methanol for extracting G-Re from American ginseng berry pulp.

Combination of Bernouilli effect producing maneuver-induced pharyngeal narrowing rate with body mass index as predictive tool for obstructive sleep apnea syndrome

Conclusions. It is suggested that the combination of the pharyngeal narrowing rate during the Bernouilli effect producing maneuver (BEPM) with the body mass index (BMI) might be a promising predictive method for obstructive sleep apnea syndrome (OSAS). Objectives. An attempt was made to clarify the possibility that the dynamic narrowing of the pharynx evaluated by nasopharyngoscopy with BEPM, a forced inspiration through the nose with the mouth closed, might be a reliable clinical daytime predictor for identifying patients at risk of developing OSAS. Subjects and methods. Subjects were 57 patients complaining of snoring and sleepiness during daytime. Endoscopic images of the retropalatal pharynx were obtained and their area was measured by NIH imaging. The pharyngeal narrowing rate during quiet nasal breathing and that during BEPM were compared to assess pharyngeal dynamics. To increase the sensitivity of this method, BMI was added to the pharyngeal narrowing rate during BEPM. Results. The criterion of BEPM at a cut-off value of 50% with BMI at a cut-off value of 25 kg/m2 achieved a sensitivity of 93%, while the specificity was 67% in differentiating simple snorers from OSAS patients (apnea index >5). In addition, the likelihood ratio of the method was found to be 2.81.

New quantitative image analysis of staphylococcal biofilms on the surfaces of nontranslucent metallic biomaterials

BackgroundImplant-related infection after orthopedic surgery is difficult to cure. One of the causes of infection is the bacterial biofilm that forms around biomaterials used during surgery. Therefore, it is necessary to investigate bacterial biofilms extensively to resolve the problems of these postoperative infections. However, no established culture method or quantification system exists for bacterial biofilms grown on the surface of the metallic biomaterials used in orthopedics, which are nonradiolucent. The purpose of this study was to develop a quantitative method to evaluate the difference in resistance of stainless steel versus titanium to staphylococcal biofilms and the efficacy of antibiotics against biofilms. MethodsThe bacterial strains used in this study were three Staphylococcus aureus stains: strain Seattle 1945 and two clinical strains cultured from postoperative infections. Staphylococcal biofilms were formed on stainless steel washers (SUS304) and titanium washers (pure titanium). They were stained with crystal violet and were examined with a digital microscope to calculate the bacterial coverage rate (BCR) by NIH imaging. ResultsThe BCR of S. aureus biofilms formed on stainless steel and titanium washers increased over time. At 24, 48, and 72h after cultivation, the amount of biofilm on the surface of the stainless steel washers was significantly greater or tended to be greater than that on the titanium. Cefazolin was applied to the obtained biofilms of two clinically isolated S. aureus strains. Cefazolin did not eradicate the biofilms but significantly reduced the biofilm of one strain. ConclusionThe newly developed quantitative method (static microtube culture and measurement system) was useful for assessing the amount of bacterial biofilms on the surface of nontranslucent biomaterial. We found that titanium may be more resistant to bacterial infection than stainless steel. To control implant-related severe infections, the biomaterials should be assessed from the viewpoint of their resistibility to bacterial adhesions and infections.

Endoscopic evaluation of dynamic narrowing of the pharynx by the Bernouilli effect producing maneuver in patients with obstructive sleep apnea syndrome

Objective To evaluate the dynamic narrowing of the retropalatal pharynx by the Bernouilli effect producing maneuver (BEPM) in patients with obstructive sleep apnea syndrome (OSAS) with computer-assisted analysis of nasopharyngoscopic images. Methods Endoscopic images of the retropalatal pharynx were obtained and their area was measured by NIH imaging. Because changes in distance between the CCD camera of the nasopharyngoscope and the site of pharyngeal narrowing decreased the measurement reliability, the area of the retropalatal pharynx was divided by the square of the uvula's lateral side length for normalization. We calculated both normalized areas: the pharyngeal narrowing during quiet nasal breathing and that during BEPM, which is a forced inspiration through the nose with the mouth closed. The narrowing rate was then calculated as an index of the pharyngeal dynamics. Results Nasopharyngoscopy combined with BEPM showed a medial motion of the lateral walls of the pharynx due to the Bernoulli effect, which is an increase in speed and a reduction in pressure of a moving gas passing through a tube constriction. There was a significant correlation between the narrowing rate of the retropalatal pharynx and the apnea index. The decrease in intraluminal pressure at the narrowing combined with the higher atmospheric pressure exercised on the lateral pharyngeal walls would induce the pharyngeal collapse. There was also a significant correlation between the normalized retropalatal are and the apnea index. Conclusion Nasopharyngoscopy with BEPM suggest that in addition to the static narrowing, the dynamic narrowing of the retropalatal pharynx contributes to the pathophysiology of OSAS. The correlation between the apnea index and both the normalized retropalatal area and the narrowing rate of the retropalatal pharynx due to the Bernoulli effect constitute a quantitative and predictive alternative to the evaluation and understanding of the upper airways changes in OSAS.

Visual detection of saikosaponins by on-membrane immunoassay and estimation of traditional Chinese medicines containing Bupleuri radix

The purpose of this study was to describe the simple, rapid, and environmental-cost effective determination method for saikosaponins in complicated samples like Bupleuri radix and traditional Chinese medicines (TCM). Saikosaponin standards, extracts of Bupleuri radix and TCM, were applied to a polyethersulphone (PES) membrane and developed by acetonitrile–water (1:4, by volume). Saikosaponin a (SSa), SSc, and SSd were visually detected by an immunostaining method (called Eastern blotting technique) using a monoclonal antibody (MAb) against SSa. At least 62.5 ng of SSa, SSc, and SSd were clearly detectable individually. These coloring spot areas of saikosaponins on PES membrane were calculated by using the NIH Imaging software and three saikosaponins can be analyzed quantitatively between 62.5 ng and 1.0 μg. Saikosaponins in Bupleuri radix and TCM were determined and these results of SSa and total saikosaponin concentrations were in good agreement with those from the ELISA analysis.

EXPERIMENTAL STUDY OF NANOPOROUS CERAMIC TUBE IN POTENTIAL APPLICATION IN HEMODIALYSIS

The self-organized nanoporous ceramic tubes offer considerable advantages over current hemodialysis membranes such as controllable pore size, uniform pore distribution, high porosity, chemical and thermal durability. The objective of this study is to fabricate and characterize nanoporous ceramic tubes and to test its hydraulic conductivity for potential use in hemodialysis. The tubular alumina membranes were anodized from an aluminum alloy in 2.7% oxalic acid at 30 V and 40 V for about 2 to 3 days. Anodization from interior to exterior made the tube strong, thereby facilitating easy mechanical handling. The membranes were imaged using a high resolution FEI SEM and characterized by using an NIH imaging software. Mini-module dialyzers of different tube lengths were made and their hydraulic conductivities were tested. The results showed that the pore size and porosity were in the range of 15.61±7.63 to 42.28±23.72 nm, and 15.77±1.86% to 49.13±4.43%, respectively, and the pore size and porosity change linearly with anodization time. The average hydraulic conductivity for the 3 cm, 6 cm and 9 cm length ceramic tubes was 1.025±0.29 x 10-15 m2Pa-1s-1, with a membrane thickness of 69 μm. This value could be increased by increasing the porosity on the outside surface of the tube using longer etching time. By using anodic alumina technology, nanoporous ceramic tubes with feature sizes approximately 15 to 45 nm can be reliably produced in quantity. These features make the nanoporous ceramic tube a potential application in hemodialysis.

ラット頭頂骨における骨形成促進剤ＴＡＫ‐７７８含有β‐ＴＣＰの効果―含有量の違いによる骨形成量の比較―

This study was conducted to histomorphologically determine the quantity of bone formation induced by TAK-778, a 3-benzothiepin derivative, (Takeda Pharmaceutical Co.Ltd.), in various amounts, using b-tricalcium phosphate granules (beta-TCP; OLYMPUS Corp.) as a carrier for the osteogenetic agent. Ten-week-old female SD rats were used. An incision was made over the parietal region of the head. The cranial periosteum was ablated and a titanium tube was fixed with an adhesive resin cement to the central part of the head, through which a mixture of TAK-778 and beta-TCP was implanted under various conditions. Tissue specimens were prepared at 4, 8, and 16 weeks after the implantation for histomorphological examination, and the proportion of new bone formation was compared at fixed time points using the NIH imaging software. The amount of new bone formation was examined every week after the implantation of TAK-778 at various doses, and the mean values were compared using Fisher's PSLD test (P< 0.05). The histomorphological observations revealed new bone formation in all the groups, irrespective of the amount and the duration of implantation of TAK-778. A comparative study revealed that the amount of new bone formation was the largest at 16 weeks following the implantation of a mixture of beta-TCP and 100 mg of TAK-778. 1. The present study confirmed the acceleration of new bone formation soon after TAK-778 implantation. 2. The results suggested that the action of TAK-778 could be maintained over time if the agent was used in combination with beta-TCP. 3. The time-course of bone formation differed depending on the proportion of TAK-778 and beta-TCP in the mixture used. 4. TAK-778 at the dose of 10 mg or 50 mg was more effective than that at the dose of 100 mg for the early formation of new bone. These results indicate that TAK-778 accelerates the formation of new bone and that beta-TCP is a useful carrier for TAK-778.

A Novel Role for Glucocorticoid-induced Leucine Zipper Protein in Epithelial Sodium Channel-mediated Sodium Transport

The steroid hormone aldosterone stimulates sodium (Na+) transport in tight epithelia by altering the expression of target genes that regulate the activity and trafficking of the epithelial sodium channel (ENaC). We performed microarray analysis to identify aldosterone-regulated transcripts in mammalian kidney epithelial cells (mpkC-CD(c14)). One target, glucocorticoid-induced leucine zipper protein (GILZ), was previously identified by serial analysis of gene expression (SAGE); however, its function in epithelial ion transport was unknown. Here we show that GILZ expression is rapidly stimulated by aldosterone in mpkCCD(c14) and that GILZ, in turn, strongly stimulates ENaC-mediated Na+ transport by inhibiting extracellular signal-regulated kinase (ERK) signaling. In Xenopus oocytes with activated ERK, heterologous GILZ expression consistently inhibited phospho-ERK expression and markedly stimulated ENaC-mediated Na+ current, in a manner similar to that of U0126 (a pharmacologic inhibitor of ERK signaling). In mpkCCD(c14) cells, GILZ transfection similarly consistently inhibited phospho-ERK expression and stimulated transepithelial Na+ transport. Furthermore, aldosterone treatment of mpkCCD(c14) cells suppressed phospho-ERK levels with a time course that paralleled their increase of Na+ transport. Finally, GILZ expression markedly increased cell surface ENaC expression in epidermal growth factor-treated mammalian kidney epithelial cells, HEK 293. These observations suggest a novel link between GILZ and regulation of epithelial sodium transport through modulation of ERK signaling and could represent an important pathway for mediating aldosterone actions in health and disease.

The NIEHS and the National Toxicology Program: An Integrated Scientific Vision

The National Toxicology Program (NTP) is an interagency program1 whose mission is to coordinate, conduct, and communicate toxicological research findings across the U.S. government. The NTP is administratively housed at the NIEHS, and David Schwartz serves as the director of both the NTP and the NIEHS. The NTP and the NIEHS share an integrated vision that serves to enhance the productivity of each program by promoting extensive collaboration across the broad spectrum of environmental health sciences. One of the emerging challenges for the NTP and the NIEHS is to use the best science to create, validate, and implement in environmental health research novel, robust, and efficient biological assays that will more effectively predict the risk of human disease and protect the health of our public. Recently the NTP developed a vision statement, “Toxicology in the 21st Century: The Role of the National Toxicology Program,” and a roadmap for implementing this vision (available at http://ntp-server.niehs.nih.gov/). The NTP vision is to develop and use the best science possible to achieve a greater understanding of the mechanisms of toxicity, and to apply this understanding to the study of a broad array of environmental agents through the most effective and efficient use of resources. This vision is consistent with the need of the NIEHS to identify and understand the human biologic and pathophysiologic response to toxicants. As we begin to develop a strategic plan for the NIEHS, an understanding of the NTP vision and roadmap can help to inform and guide this process. The NTP roadmap has identified four key scientific areas as priorities. First, the NTP needs to modernize the mammalian assays used to screen for toxicity. NIEHS research will lead the development of such assays. Critical to this process is increasing our understanding of the similarities and differences between laboratory species and humans. It is clear that both the quality and the interpretability of toxicity data will improve through the strategic use of new approaches to comparative biology. Second, the NTP needs to develop and implement high- and medium-throughput screens to identify and understand potential targets for environmentally mediated disease. Such screens, ranging in complexity from simple subcellular fractions to complicated mixtures of primary cultures, can address a variety of biochemical, mechanistic, and functional end points. The availability of these screens will allow the NTP to establish priorities for full-scale, resource-intensive mammalian assays, and will provide direct links into the hypothesis-driven research supported by the NIEHS. As part of this process, the NTP and the NIEHS are collaborating with investigators in the NIH Roadmap Molecular Libraries and Imaging Initiative who are screening more than 100,000 compounds against multiple cellular targets to identify possible therapeutic agents and basic biologic responses. The in vivo toxicity data in the NTP archives will be incorporated into this high-throughput screen, and will serve as a cornerstone for the NIEHS and others to develop linkages between basic biologic responses and pathophysiologic outcomes. Third, recent developments in biomedical research and molecular genetics have created a tremendous need to develop better ways to store, retrieve, analyze, and interpret vast amounts of data. The need for databases and repositories is critical for evaluating the toxicity of potentially hazardous agents. The NTP and the NIEHS will play important and complementary roles in developing these resources, and will partner with others to develop similar tools for the wider range of toxicologic, biologic, genetic, genomic, and biochemical information. Finally, training the next generation of scientists is critical to the environmental health sciences. In collaboration with the NTP member agencies and our colleagues at the National Institutes of Health, we will support the development of training programs focused on creating integrated teams of scientists to understand and attack environmental health problems of concern to the public. The NTP and the NIEHS share an integrated vision that serves to enhance the productivity of each program by promoting extensive collaboration across the broad spectrum of environmental health sciences. The fields of toxicology and environmental health sciences are intimately linked, and the future for both is challenging. We are excited by the possibilities posed by the new directions of the NTP and the NIEHS, and look forward to the continued evolution of this vital and productive relationship.

Parkin Stabilizes Microtubules through Strong Binding Mediated by Three Independent Domains

Mutations of parkin, a protein-ubiquitin isopeptide ligase (E3), appear to be the most frequent cause of familial Parkinson's disease (PD). Our previous studies have demonstrated that parkin binds strongly to alpha/beta tubulin heterodimers and microtubules. Here we show that the strong binding between parkin and tubulin, as well as that between parkin and microtubules, was mediated by three independent domains: linker, RING1, and RING2. These redundant strong interactions made it virtually impossible to separate parkin from microtubules by high concentrations of salt (3.8 m) or urea (0.5 m). Parkin co-purified with tubulin and was found in highly purified tubulin preparation. Expression of either full-length parkin or any of its three microtubule-binding domains significantly attenuated colchicine-induced microtubule depolymerization. The abilities of parkin to bind to and stabilize microtubules were not affected by PD-linked mutations that abrogate its E3 ligase activity. Thus, the tubulin/microtubule-binding activity of parkin and its E3 ligase activity are independent. The strong binding between parkin and tubulin/microtubules through three redundant interaction domains may not only stabilize microtubules but also guarantee the anchorage of this E3 ligase on microtubules. Because many misfolded proteins are transported on microtubules, the localization of parkin on microtubules may provide an important environment for its E3 ligase activity toward misfolded substrates.

マウスの酢酸に対する忌避行動の解析

Gene alignments of mice have been well-identified, and many transgenic mice have been studied to determine mechanisms behind diseases. Some transgenic mice have been reported to show olfactory dysfunction, and they are expected to increase in number. The conventional analysis of olfactory behavior is complicated and takes much time. We report a new trial for analysis of olfactory behavior using video cameras and NIH imaging. We observed that mice avoided cotton balls including acetic acid placed in a corner of a square cage, and recorded that behavior with video cameras. Motion pictures were loaded onto a personal computer and analyzed using NIH imaging. The cage was divided into 4 areas and the time mice remained in each area was calculated. Mice were evenly distributed in areas without the cotton ball with acetic acid. They remained significantly shorter in area with the cotton ball. Results suggest that this new analys may be useful for studying olfactory behavior in mice, though additional studies are required.

骨形成促進剤ＴＡＫ‐７７８‐ＳＲの担体に用いたβ‐ｔｒｉｃａｌｃｉｕｍ ｐｈｏｓｐｈａｔｅの評価

The aim of this study was to evaluate the combination of TAK-778-SR which was sustained-release microcapsules of a bone formation stimulant, TAK-778, and its carrier beta-tricalcium phosphate (beta-TCP) blocks (pore rates 60%, 75% respectively). The difference of their abilities in bone-formation was evaluated histomorphologically by varying the following conditions: with or without TAK-778, pore ratio of carrier and embedding period. Nine-week-old female SD rats were used. After removing the parietal bone from the head with a trephine bar, the defects were refilled by beta-TCP blocks immersed with or without TAK-778 under the following conditions: saline solution, release microcapsules only, and release microcapsules with TAK-778 (TAK-778-SR). These rats were sacrificed after 5 and 10 weeks and their histological specimens were prepared. Morphological change was observed and the formation rates of each new bone were compared using an NIH imaging program. A significant amount of new bone was morphologically observed in all beta-TCP samples that were treated with TAK-778-SR. A high rate of new bone formation was confirmed in the 10-week samples (pore rate 75%, with TAK-778-SR) with the NIH imaging. 1. beta-TCP and release microcapsules did not disturb the recovery process. 2. Five- and 10-week samples (pore rate 60%) were absorbed marginally. 3. Absorption was observed in the 5-week samples (pore rate 75%), and it was accelerated further at 10 weeks. 4. An accelerating bone-formation effect of TAK-778-SR was confirmed and beta-TCP block was proved to be highly useful as a carriage material.

965. A Novel Approach for Gene Transduction with Adenovirus/Fibrin Glue System

Introduction: The major cause of local recurrence after initial surgery for hepatic malignancy is residual carcinoma macroscopically or microscopically at the surgical site. Although many surgical devices and skills have developed, the recurrence rate of hepatic malignancy is still high. At present, for those patients with recurrent disease, adjuvant therapies such as chemotherapy, radiation therapy and immunotherapy are combined. However, in such cases, severe side-effects make the patient's QOL (quality of life) considerably worse. Therefore, a novel approach for residual carcinoma at the time of surgery is urgently needed. In this study we proposed the use of a novel gene therapy for residual carcinoma of the liver. Purpose: The fibrin glue system (FGS) consists of liquid forms of fibrinogen and thrombin and is used widely in surgical operation as it has a sealing effect and promotes tissue adhesion at the surgically resected site. In this study, we proposed the efficacy and safety of loco-regional gene transfer using adenoviral vector mixed with FGS to prevent local recurrence of liver carcinoma. We demonstrated that the adenoviral vector containing the E.coli LacZ gene (AxCALacZ) was delivered to the liver of rats by FGS. Materials and Methods: Experiment 1 in vitro Rat colon carcinoma cell line, RCN-9 was incubated on a membrane containing multiple 0.02μm-sized holes. Various concentrations of the adenoviral vector mixed with FGS (AxCALacZ/FGS) were applied on the RCN-9 and incubated for 48 hours. The optimum efficiency of transduction was evaluated by X-Gal staining and NIH imaging. Experiment 2 in vitro To examine the survival period of adenoviral vector in FGS, AxCALacZ/FGS put on the monolayer cultured RCN9 after being left for various periods. Transgene expression was evaluated by β-gal assay. The β-gal activity was measured photometrically using the Promega β-gal enzyme assay kit according to the manufacture's instructions (Promega, Madison, USA). Experiment 3 in vivo 8-week-old male F344 rats underwent a partial hepatectomy. AxCALacZ/FGS was applied on the cut surface of the dissected liver and the rats were kept for 48 hours under normal conditions. AxLacZ/PBS was treated as the control study. The transducted and non-transducted parts of the liver were extirpated, stained with X-gal liquid and β-gal was measured to compare the efficacy of transduction in each organ. Results: Experiment 1 significantly more transgene expression was shown by the X-gal staining and NIH imaging than 1MOI. Experiment 2 LacZ expression revealed infected RCN9 when AxCALacZ/FGS was left for a period of less than 96 hours. Experiment 3 AxCALacZ/FGS transgene expression was shown at the treated and non-treated part of the liver more than AxCALacZ/PBS expression. Discussion and conclusion: It was demonstrated that adenoviral vector survives and being stable for enough time to transduct in FGS. AxCALacZ/FGS can transduct the target gene. Furthermore, the transduction efficiency was comparable to those of existing methods. In the future, FGS will be studied with an adenoviral vector containing the gene which enhances antitumor immunity for preventing recurrence loco-regionally site as well as its original tissue adhesive purpose.

Tyrosine phosphorylation of QKI mediates developmental signals to regulate mRNA metabolism.

The selective RNA-binding protein QKI is essential for myelination in the central nervous system (CNS). QKI belongs to the family of signal transduction activators of RNA (STARs), characteristic of binding RNA and signaling molecules, therefore is postulated to regulate RNA homeostasis in response to developmental signals. Here we report that QKI acts downstream of the Src family protein tyrosine kinases (Src-PTKs) during CNS myelination. QKI selectively interacted with the mRNA encoding the myelin basic protein (MBP). Such interaction stabilized MBP mRNA and was required for the rapid accumulation of MBP mRNA during active myelinogenesis. We found that the interaction between QKI and MBP mRNA was negatively regulated by Src-PTK-dependent phosphorylation of QKI. During early myelin development, tyrosine phosphorylation of QKI in the developing myelin drastically declined, presumably leading to enhanced interactions between QKI and MBP mRNA, which was associated with the rapid accumulation of MBP mRNA and accelerated myelin production. Therefore, developmental regulation of Src-PTK-dependent tyrosine phosphorylation of QKI suggests a novel mechanism for accelerating CNS myelinogenesis via regulating mRNA metabolism.

Reduced p21, p27 and vitamin D receptor in the nodular hyperplasia in patients with advanced secondary hyperparathyroidism1

Reduced p21, p27 and vitamin D receptor in the nodular hyperplasia in patients with advanced secondary hyperparathyroidism.BackgroundIn uremic patients with secondary hyperparathyroidism (2HPT), nodular hyperplasia of parathyroid gland shows a monoclonal pattern of cell proliferation, in which a decreased density of vitamin D receptor (VDR) also is demonstrated. The present study aimed at elucidating the mechanism of parathyroid cell proliferation in relation to cell cycle determinants in patients with advanced 2HPT.MethodsThe expression of cyclin-dependent kinase inhibitors, p21 and p27, and VDR were examined and compared among four groups of nodular (Nd; N = 23) or diffuse (Df; N = 6) hyperplastic parathyroid glands resected due to 2HPT, primary adenomas (Ad; N = 15), and histologically-normal parathyroid glands (C; N = 20) removed during thyroidectomy. Immunohistochemical analyses for VDR, p21, p27 and Ki67 antigen were performed in formalin-fixed paraffin-embedded tissues by using specific polyclonal antibody. The distribution and the intensity of immunoreactivity was quantified by using NIH imaging, and was expressed as the labeling index (LI) of positive nuclear staining in a random set of 1000 cells.Resultsp21 LI was significantly diminished in both Nd (85 ± 110; mean ± SD) and Ad (136 ± 122) as compared to that in Df (360 ± 191) or C (359 ± 228; P < 0.01). p27 LI was also significantly diminished in both Nd (97 ± 156) and Ad (187 ± 196) as compared to that in Df (532 ± 146) or C (631 ± 170; P < 0.01). VDR LI in Nd (162 ± 194) was also significantly lower than that in Df (495 ± 337), Ad (383 ± 262), or C (659 ± 234), respectively (P < 0.01). Parathyroid sections with high nuclear VDR expression elicited high p21 and p27 expression. Both p21 and p27 LI in Nd correlated significantly with nuclear VDR LI (r = 0.92; P < 0.01, r = 0.76; P < 0.01), but not with p53 LI, and inversely correlated with the glandular weight (r = 0.44; P < 0.05, r = 0.41; P < 0.05).ConclusionsThe reduced expression of p21 and p27, in a VDR-dependent manner, is a major pathogenic factor for a nodular parathyroid gland growth. Reduced p21, p27 and vitamin D receptor in the nodular hyperplasia in patients with advanced secondary hyperparathyroidism. In uremic patients with secondary hyperparathyroidism (2HPT), nodular hyperplasia of parathyroid gland shows a monoclonal pattern of cell proliferation, in which a decreased density of vitamin D receptor (VDR) also is demonstrated. The present study aimed at elucidating the mechanism of parathyroid cell proliferation in relation to cell cycle determinants in patients with advanced 2HPT. The expression of cyclin-dependent kinase inhibitors, p21 and p27, and VDR were examined and compared among four groups of nodular (Nd; N = 23) or diffuse (Df; N = 6) hyperplastic parathyroid glands resected due to 2HPT, primary adenomas (Ad; N = 15), and histologically-normal parathyroid glands (C; N = 20) removed during thyroidectomy. Immunohistochemical analyses for VDR, p21, p27 and Ki67 antigen were performed in formalin-fixed paraffin-embedded tissues by using specific polyclonal antibody. The distribution and the intensity of immunoreactivity was quantified by using NIH imaging, and was expressed as the labeling index (LI) of positive nuclear staining in a random set of 1000 cells. p21 LI was significantly diminished in both Nd (85 ± 110; mean ± SD) and Ad (136 ± 122) as compared to that in Df (360 ± 191) or C (359 ± 228; P < 0.01). p27 LI was also significantly diminished in both Nd (97 ± 156) and Ad (187 ± 196) as compared to that in Df (532 ± 146) or C (631 ± 170; P < 0.01). VDR LI in Nd (162 ± 194) was also significantly lower than that in Df (495 ± 337), Ad (383 ± 262), or C (659 ± 234), respectively (P < 0.01). Parathyroid sections with high nuclear VDR expression elicited high p21 and p27 expression. Both p21 and p27 LI in Nd correlated significantly with nuclear VDR LI (r = 0.92; P < 0.01, r = 0.76; P < 0.01), but not with p53 LI, and inversely correlated with the glandular weight (r = 0.44; P < 0.05, r = 0.41; P < 0.05). The reduced expression of p21 and p27, in a VDR-dependent manner, is a major pathogenic factor for a nodular parathyroid gland growth.

Reduced p21, p27 and vitamin D receptor in the nodular hyperplasia in patients with advanced secondary hyperparathyroidism

In uremic patients with secondary hyperparathyroidism (2HPT), nodular hyperplasia of parathyroid gland shows a monoclonal pattern of cell proliferation, in which a decreased density of vitamin D receptor (VDR) also is demonstrated. The present study aimed at elucidating the mechanism of parathyroid cell proliferation in relation to cell cycle determinants in patients with advanced 2HPT. The expression of cyclin-dependent kinase inhibitors, p21 and p27, and VDR were examined and compared among four groups of nodular (Nd; N = 23) or diffuse (Df; N = 6) hyperplastic parathyroid glands resected due to 2HPT, primary adenomas (Ad; N = 15), and histologically-normal parathyroid glands (C; N = 20) removed during thyroidectomy. Immunohistochemical analyses for VDR, p21, p27 and Ki67 antigen were performed in formalin-fixed paraffin-embedded tissues by using specific polyclonal antibody. The distribution and the intensity of immunoreactivity was quantified by using NIH imaging, and was expressed as the labeling index (LI) of positive nuclear staining in a random set of 1000 cells. p21 LI was significantly diminished in both Nd (85 +/- 110; mean +/- SD) and Ad (136 +/- 122) as compared to that in Df (360 +/- 191) or C (359 +/- 228; P < 0.01). p27 LI was also significantly diminished in both Nd (97 +/- 156) and Ad (187 +/- 196) as compared to that in Df (532 +/- 146) or C (631 +/- 170; P < 0.01). VDR LI in Nd (162 +/- 194) was also significantly lower than that in Df (495 +/- 337), Ad (383 +/- 262), or C (659 +/- 234), respectively (P < 0.01). Parathyroid sections with high nuclear VDR expression elicited high p21 and p27 expression. Both p21 and p27 LI in Nd correlated significantly with nuclear VDR LI (r = 0.92; P < 0.01, r = 0.76; P < 0.01), but not with p53 LI, and inversely correlated with the glandular weight (r = 0.44; P < 0.05, r = 0.41; P < 0.05). The reduced expression of p21 and p27, in a VDR-dependent manner, is a major pathogenic factor for a nodular parathyroid gland growth.

REDUCTION IN PATELLOFEMORAL PAIN IS ASSOCIATED WITH INCREASES IN CONTACT AREA FOLLOWING PATELLAR BRACING

Elevated patellofemoral joint stress (force/unit area) as a result of patellar malalignment has been hypothesized as being contributory to patellofemoral pain. Clinically, bracing has been reported to decrease patellofemoral pain, however studies have shown minimal or no changes in patellar alignment. It is our hypothesis that decreases in pain are associated with increases in patellofemoral contact area, as forces would be distributed over a larger surface area. PURPOSE: To evaluate the relationship between changes in patellofemoral pain and changes in patellofemoral joint contact area. METHODS: Seven females (age: 31 ± 8y; height: 165 ± 9cm; body mass: 56 ± 6kg) with readily reproducible pain participated in this study. Subjects were first asked to rate their pain during a unilateral squat using a visual analog scale. A patellofemoral joint brace was then applied and the unilateral squat and visual analog scale pain rating was repeated. Following the pain assessment, magnetic resonance imaging was conducted while performing isometric contractions against 25% body weight at 0, 20, 40, & 60 degrees of knee flexion. This was repeated for both braced and non-braced conditions. Using a previously established protocol, measurements of patellofemoral joint contact area were made using NIH imaging software. Pearson's correlation coefficients were used to quantify the relationship between changes in contact area and changes in patellofemoral pain. RESULTS: On the average, subjects reported a 40% decrease in pain following bracing. Significant increases in contact area were observed in the braced condition across all knee flexion angles evaluated. Decreases in patellofemoral pain were found to be significantly associated with increases in contact area at 0° (r = .42), 20° (r = .69), and at 40° (r = .56.) CONCLUSION: These results indicate that increases in contact area may contribute to decreased symptoms in persons with patellofemoral pain. The compression imparted to the patellofemoral joint as a result of bracing likely seats the patella more firmly in the trochlear groove, thereby distributing joint reaction forces over a greater surface area (ie. decreased stress.) These findings support the use of patellofemoral bracing as an intervention to reduce patellofemoral pain. Supported by the unrestricted grant from Don Joy, Inc.

Kupffer Cell Inactivation Delays Repair in a Rat Model of Reversible Biliary Obstruction

During cholestatic liver injury, Kupffer cells (KC) and activated macrophages modulate cell proliferation and subsequent matrix deposition. The role of KC in the restoration of cell architecture and matrix metabolism during repair following chronic cholestatic liver injury is unknown. To determine the effect of KC inactivation, adult male Sprague-Dawley rats underwent bile duct suspension (BDS) for 5 days followed by reversal of the obstruction. Saline (control) and gadolinium chloride (10 mg/kg) were administered 1 day prior to BDS and 1 day prior to reversal, to inactivate KC during both injury and repair. Serum bilirubin and quantitative cell morphometry were compared to verify the reversibility of the model. Collagen content of the liver was measured in trichrome-stained paraffin sections using NIH imaging software. Reversibility of the obstruction was verified by normalization of direct serum bilirubin, which peaked at 8.42 +/- 0.76 mg/dL following 5 days of BDS and returned to sham-operated levels 2 days after reversal, 0.36 +/- 0.15 mg/dL. Hematoxylin and eosin (H&E)-stained paraffin-embedded liver sections from gadolinium-treated animals at 4 and 7 days after reversal exhibited persistent bile duct proliferation, matrix deposition, and inflammation. Gadolinium-treated animals had altered collagen metabolism compared to saline controls. Whereas the collagen content in the saline group slowly returned to sham-operated levels over time, the treatment group demonstrated progressive accumulation of collagen during repair which was statistically significant at 7 days following reversal (8.79%/mm(2) +/- 2.17 in gadolinium group vs 2. 33%/mm(2) +/- 0.34 in saline group, P = 0.0003). These results demonstrate that inactivation of resident hepatic macrophages during liver repair impairs collagen metabolism, inhibits the resolution of fibrosis, and allows the persistence of inflammatory cell infiltrates in the portal areas. This is the first evidence of profibrogenic responses in the absence of an intact KC compartment during repair after cholestatic injury.