Molecular Enhancement of Alpha 7 Integrin to Ameliorate Muscular Dystrophy

126 p.Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2007.Duchenne muscular dystrophy is the most common form of muscular dystrophy and it is lethal; most patients dye before their early twenties. This disease is caused by mutations in the gene encoding dystrophin, a member of the dystrophin protein complex that links the extracellular matrix to the cytoskeleton in skeletal muscle. The alpha7beta1 integrin also links laminin in the extracellular matrix to actin in skeletal muscle. It has been shown that transgenic overexpression of the alpha7 integrin chain in mdx/utr (-/-) mice, a model for Duchenne muscular dystrophy, ameliorates dystrophic pathology and prolongs survival. To translate this result into a therapy for patients, myogenic cells from the alpha7(+/-) mice that have the beta galactosidase reporter driven by the alpha7 integrin promoter were screened for molecules that enhance alpha7 expression. Two such molecules were identified: HTCB and valproic acid increase alpha7 levels about 1.4-fold and 2-fold respectively. Valproic acid also activates the Akt/mTOR pathway, promoting myotube hypertrophy and survival. To test their effects in vivo, the molecules were administered to mdx/utr (-/-) mice. Valproic acid injected mice have decreased fibrosis and contractures, improved mobility and kyphosis, decreased inflammatory cell content and cardiomyopathy and increased myofiber integrity. HTCB injected mice have increased myofiber integrity and show elevated alpha7 integrin expression in muscle. Together, these results represent important milestones towards developing an alpha7beta1 integrin based complementary gene therapy for Duchenne muscular dystrophy.U of I OnlyRestricted to the U of I community idenfinitely during batch ingest of legacy ETD

Mobile microscopy as a screening tool for oral cancer in India: A pilot study.

Oral cancer is the most common type of cancer among men in India and other countries in South Asia. Late diagnosis contributes significantly to this mortality, highlighting the need for effective and specific point-of-care diagnostic tools. The same regions with high prevalence of oral cancer have seen extensive growth in mobile phone infrastructure, which enables widespread access to telemedicine services. In this work, we describe the evaluation of an automated tablet-based mobile microscope as an adjunct for telemedicine-based oral cancer screening in India. Brush biopsy, a minimally invasive sampling technique was combined with a simplified staining protocol and a tablet-based mobile microscope to facilitate local collection of digital images and remote evaluation of the images by clinicians. The tablet-based mobile microscope (CellScope device) combines an iPad Mini with collection optics, LED illumination and Bluetooth-controlled motors to scan a slide specimen and capture high-resolution images of stained brush biopsy samples. Researchers at the Mazumdar Shaw Medical Foundation (MSMF) in Bangalore, India used the instrument to collect and send randomly selected images of each slide for telepathology review. Evaluation of the concordance between gold standard histology, conventional microscopy cytology, and remote pathologist review of the images was performed as part of a pilot study of mobile microscopy as a screening tool for oral cancer. Results indicated that the instrument successfully collected images of sufficient quality to enable remote diagnoses that show concordance with existing techniques. Further studies will evaluate the effectiveness of oral cancer screening with mobile microscopy by minimally trained technicians in low-resource settings

A smart tele-cytology point-of-care platform for oral cancer screening.

Early detection of oral cancer necessitates a minimally invasive, tissue-specific diagnostic tool that facilitates screening/surveillance. Brush biopsy, though minimally invasive, demands skilled cyto-pathologist expertise. In this study, we explored the clinical utility/efficacy of a tele-cytology system in combination with Artificial Neural Network (ANN) based risk-stratification model for early detection of oral potentially malignant (OPML)/malignant lesion. A portable, automated tablet-based tele-cytology platform capable of digitization of cytology slides was evaluated for its efficacy in the detection of OPML/malignant lesions (n = 82) in comparison with conventional cytology and histology. Then, an image pre-processing algorithm was established to segregate cells, ANN was trained with images (n = 11,981) and a risk-stratification model developed. The specificity, sensitivity and accuracy of platform/ stratification model were computed, and agreement was examined using Kappa statistics. The tele-cytology platform, Cellscope, showed an overall accuracy of 84-86% with no difference between tele-cytology and conventional cytology in detection of oral lesions (kappa, 0.67-0.72). However, OPML could be detected with low sensitivity (18%) in accordance with the limitations of conventional cytology. The integration of image processing and development of an ANN-based risk stratification model improved the detection sensitivity of malignant lesions (93%) and high grade OPML (73%), thereby increasing the overall accuracy by 30%. Tele-cytology integrated with the risk stratification model, a novel strategy established in this study, can be an invaluable Point-of-Care (PoC) tool for early detection/screening in oral cancer. This study hence establishes the applicability of tele-cytology for accurate, remote diagnosis and use of automated ANN-based analysis in improving its efficacy

Valproic Acid Activates the PI3K/Akt/mTOR Pathway in Muscle and Ameliorates Pathology in a Mouse Model of Duchenne Muscular Dystrophy

Duchenne muscular dystrophy is a lethal neuromuscular disease that currently has no effective therapy. Transgenic overexpression of the α7 integrin in mdx/utrn−/− mice, a model of Duchenne muscular dystrophy ameliorates the disease. We have isolated and used α7+/− muscle cells expressing β-galactosidase, driven by the endogenous α7 promoter, to identify compounds that increase α7 integrin levels. Valproic acid (VPA) was found to enhance α7 integrin levels, induce muscle hypertrophy, and inhibit apoptosis in myotubes by activating the Akt/mTOR/p70S6K pathway. This activation of the Akt pathway occurs within 1 hour of treatment and is mediated by phosphatidylinositol 3-OH kinase. To evaluate the potential use of VPA to treat muscular dystrophy, mdx/utrn−/− mice were injected with the drug. Treatment with VPA lowered collagen content and fibrosis, and decreased hind limb contractures. VPA-treated mice also had increased sarcolemmal integrity and decreased damage, decreased CD8-positive inflammatory cells, and higher levels of activated Akt in their muscles. Thus, VPA has important biological effects that may be applicable for the treatment of muscular dystrophy

Workflow of sample acquisition and analysis for the study.

<p>Workflow of sample acquisition and analysis for the study.</p

Sensitivity and specificity of the automated CellScope vs. histology.

<p>Sensitivity and specificity of the automated CellScope vs. histology.</p

Screenshots of the CellScope server user interface.

<p>(A) Screenshot of the CellScope server (web portal interface) showing the list of patients whose diagnosis has been carried out. Color-coding of records was used to indicate the status of the samples (i.e. diagnostic result provided, awaiting review, sample rejected, etc.) (B) Screenshot of server interface showing CellScope images acquired from a patient sample. Note the thumbnail panel on the left showing thumbnails of all images captured for that patient sample and a magnified image of the selected image in the center/right. Selected regions of interest (indicated by light blue rectangles which could be drawn by the pathologists using tools provided in the interface) are overlaid on the image. (C) Screenshot of the server interface showing a pop-up window containing various cellular features which could be used by the pathologist to annotate the selected region of interest. A free-text box in this window allowed the pathologists to enter additional comments, if necessary.</p

Sensitivity and specificity of the automated CellScope vs. cytology.

<p>Sensitivity and specificity of the automated CellScope vs. cytology.</p