Modeling non-clinical and clinical drug tests in Gaucher disease.

There is need for modeling biological systems to accelerate drug pipelines for treating metabolic diseases. The eliglustat treatment for Gaucher disease is approved by the FDA with a companion genomic test. The Transcriptome-To-Metabolome™ biosimulation technology was used to model, in silico, a standard non-clinical eliglustat test with an in vitro canine kidney cell system over-expressing a human gene; and a clinical test using human fibroblasts from control and Gaucher disease subjects.

Macrophage TGF-1 and the Proapoptotic Extracellular Matrix Protein BIGH3 Induce Renal Cell Apoptosis in Prediabetic and Diabetic Conditions.

Metabolically stressed kidney is in part characterized by infiltrating macrophages and macrophage-derived TGF-1 that promote the synthesis of various ECM molecules. TGF-1 strongly enhances the expression of the gene that encodes a cell-adhesion class, proapoptotic ECM protein called BIGH3. We hypothesized that in a diabetic environment a relationship between infiltrating macrophages, macrophage-derived TGF-1, and BIGH3 protein promotes renal cell death.

TGFβ induces BIGH3 expression and human retinal pericyte apoptosis: a novel pathway of diabetic retinopathy.

PurposeOne of the earliest hallmarks of diabetic retinopathy is the loss of retinal pericytes. However, the mechanisms that promote pericyte dropout are unknown. In the present study, we propose a novel pathway in which pericyte apoptosis is mediated by macrophages, TGFβ and pro-apoptotic BIGH3 (TGFβ-induced Gene Human Clone 3) protein.

Biomarkers from biosimulations: Transcriptome-To-Reactome™ Technology for individualized medicine.

We validated a model of the TGF-β signaling pathway using reactions from Reactome. Using a patentpending technique, gene expression profiles from individual patients are used to determine model parameters. Gene expression profiles from 45 women, normal, or benign tumor and malignant breast cancer were used as training and validating sets for assessing clinical sensitivity and specificity.

BIGH3 protein and macrophages in retinal endothelial cell apoptosis.

Diabetes is a pandemic disease with a higher occurrence in minority populations. The molecular mechanism to initiate diabetes-associated retinal angiogenesis remains largely unknown. We propose an inflammatory pathway of diabetic retinopathy in which macrophages in the diabetic eye provide TGFβ to retinal endothelial cells (REC) in the retinal microvasculature.

Transcriptional regulation of proteoglycan 4 by 17β-estradiol in immortalized baboon temporomandibular joint disc cells.

Temporomandibular joint disorders (TMDs) affect a significant portion of the population of the USA, with the majority of those seeking treatment being women of childbearing age. Owing to this striking sexual dimorphism it has been postulated that sex hormones play a role in the maintenance of normal temporomandibular joint (TMJ) function. Proteoglycan 4 (PRG4) is a secreted lubricating molecule required for maintaining low frictional levels within articular joints; however, its role in the TMJ is not well characterized.

Proteomic insights into the protective mechanisms of an in vitro oxidative stress model of early stage Parkinson's disease.

Previous studies in Parkinson's disease (PD) models suggest that early events along the path to neurodegeneration involve activation of the ubiquitin-proteasome system (UPS), endoplasmic reticulum-associated degradation (ERAD), and the unfolded protein response (UPR) pathways, in both the sporadic and familial forms of the disease, and thus ER stress may be a common feature. Furthermore, impairments in protein degradation have been linked to oxidative stress as well as pathways associated with ER stress. We hypothesize that oxidative stress is a primary initiator in a multi-factorial cascade driving dopaminergic (DA) neurons towards death in the early stages of the disease.

Up-regulation of stromal versican expression in advanced stage serous ovarian cancer.

Objective: The purpose of this study is to examine the role of versican (VCAN) in advanced stage serous ovarian cancer by investigating its expression, its function, and its correlation with clinical outcomes.

Methods: Microarray analysis was performed on RNA isolated from tumor and stromal components of advanced stage serous ovarian cancer and normal ovarian epithelial tissue to identify genes up-regulated in ovarian tumor stroma. Validation studies using immunohistochemistry and quantitative real-time PCR (Q-RT-PCR) was performed on one of the up-regulated genes, VCAN.

Thiol oxidative stress induced by metabolic disorders amplifies macrophage chemotactic responses and accelerates atherogenesis and kidney injury in LDL receptor-deficient mice.

Background: Strengthening the macrophage glutathione redox buffer reduces macrophage content and decreases the severity of atherosclerotic lesions in LDL receptor-deficient (LDLR(-/-)) mice, but the underlying mechanisms were not clear. This study examined the effect of metabolic stress on the thiol redox state, chemotactic activity in vivo, and the recruitment of macrophages into atherosclerotic lesions and kidneys of LDL-R(-/-) mice in response to mild, moderate, and severe metabolic stress.

Methods And Results: Reduced glutathione (GSH) and glutathione disulfide (GSSG) levels in peritoneal macrophages isolated from mildly, moderately, and severe metabolically-stressed LDL-R(-/-) mice were measured by HPLC, and the glutathione reduction potential (E(h)) was calculated.

C-terminal fragment of transforming growth factor beta-induced protein (TGFBIp) is required for apoptosis in human osteosarcoma cells.

Transforming growth factor beta-induced protein (TGFBIp), is secreted into the extracellular space. When fragmentation of C-terminal portions is blocked, apoptosis is low, even when the protein is overexpressed. If fragmentation occurs, apoptosis is observed.

The accumulation of versican in the nodules of benign prostatic hyperplasia.

Background: Proteoglycans, a complex group of extracellular matrix (ECM) molecules, are elevated in benign prostatic hyperplasia (BPH). Versican is a stromal proteoglycan present in prostate tissue. Versican expression is elevated in tissues with increased proliferation.

Prevalence of diabetes mellitus and correlation of urinary transforming growth factor-beta1 with blood hemoglobin A1C in the Atascosa Diabetes Study.

Introduction: This study was conducted to determine the prevalence of type 2 diabetes and prediabetes in the Atascosa Diabetes Study sample and to ascertain the relationship between urinary transforming growth factor-beta1 (TGF-beta1) and blood hemoglobin (Hgb) A1C.

Methods: Subjects (N = 526) classified as adjusted normal, at risk, prediabetes, and diabetes mellitus were given a one-hour and two-hour postprandial glucose (PPG) test. Morning urine samples were collected to test for a correlation of TGF-beta1 with blood HgbA1C.

A focal adhesion-like process underlies induction of long-term potentiation in the Schaffer collateral-CA1 region of the hippocampus.

In the series of experiments reported here we provide evidence that a focal adhesion-like process underlies the induction of long-term potentiation (LTP) in the Schaffer Collateral-CA1 projection in the hippocampus. Here we show that an integrin binding peptide (RGD) impairs induction of Schaffer Collateral-CA1 LTP in hippocampal slice preparations in vitro. The heparin-binding peptide that binds heparan sulfate proteoglycan (HSPG) and blocks the formation of focal adhesions also impairs induction of LTP.

Interleukin-1beta selectively decreases the synthesis of versican by arterial smooth muscle cells.

Proteoglycans accumulate in lesions of atherosclerosis but little is known as to which factors regulate the synthesis of these molecules. Interleukin-1beta (IL-1beta) is a cytokine involved in vascular lesion development but it is not clear whether it has specific effects on proteoglycan synthesis by arterial smooth muscle cells (ASMC). Monkey ASMC were treated with IL-1beta and proteoglycan synthesis assessed using [(35)S]-sulfate and [(35)S]-Trans amino acid labeling.

Differences in the magnitude of long-term potentiation produced by theta burst and high frequency stimulation protocols matched in stimulus number.

Theta-burst stimulation (TBS: four pulses at 100 Hz repeated with 200 ms inter-burst-intervals) and another commonly used high-frequency stimulation protocol (HFS: 1 s burst of equally spaced pulses at 100 Hz) were compared for the magnitude of LTP produced in rat hippocampal slices. The total number of pulses applied during tetanus (TET) was either 40, 100, 200, or 300. In a conventional analysis of the last 10 min of the post-TET period, a two-way ANOVA revealed no difference either in LTP of the field excitatory post-synaptic potential (fEPSP) between TBS and HFS or differences across pulse number at 40, 100, or 200 pulses.

Regulated expression of ADAMTS family members in follicles and cumulus oocyte complexes: evidence for specific and redundant patterns during ovulation.

Protease cascades are essential for many biological events, including the LH-induced process of ovulation. ADAMTS1 (a disintegrin and metalloproteinase with thrombospondin-like repeats-1) is expressed and hormonally regulated in the ovary by LH and the progesterone receptor. To determine whether other family members might be expressed and regulated in the rodent ovary, those closely related to ADAMTS1 (ADAMTS4 and ADAMTS5) were analyzed in the mouse ovary by reverse transcription-polymerase chain reaction as well as by Western blot, immunohistochemical, and immunocytochemical analyses using highly specific antibodies.

In vitro osteogenic differentiation of marrow stromal cells encapsulated in biodegradable hydrogels.

Novel hydrogel materials based on oligo(poly(ethylene glycol) fumarate) (OPF) crosslinked with a redox radical initiation system were recently developed in our laboratory as injectable cell carriers for orthopedic tissue engineering applications. The effect of OPF hydrogel material properties on in vitro osteogenic differentiation of encapsulated rat marrow stromal cells (MSCs) with and without the presence of osteogenic supplements (dexamethasone) was investigated. Two OPF formulations that resulted in hydrogels with different swelling properties were used to encapsulate rat MSCs (seeding density approximately 13 million cells/mL, samples 6 mm diameter x 0.

Expression of extracellular matrix components versican, chondroitin sulfate, tenascin, and hyaluronan, and their association with disease outcome in node-negative breast cancer.

Purpose: The purpose is to determine whether the levels of expression of extracellular matrix components in peritumoral stroma are predictive of disease outcome for women with node-negative breast cancer.

Experimental Design: Tumor tissue from 86 patients with node-negative breast cancer was examined by immunohistochemical staining for the expression of versican, chondroitin sulfate (CS), tenascin, and hyaluronan (HA). With the exception of HA, the expression of the extracellular matrix components was measured by video image analysis.

Developmental expression patterns of Beta-ig (betaIG-H3) and its function as a cell adhesion protein.

Beta-ig is a secretory protein embodied by fasciclin I-like repeats containing sequences that might bind integrins and glycosaminoglycans in vivo. Expression of Beta-ig is responsive to Transforming Growth Factor-beta and the protein is found to be associated with extracellular matrix (ECM) molecules, implicating Beta-ig as an ECM adhesive protein of developmental processes. The spatiotemporal distribution of Beta-ig during various stages of murine development was examined and its ability to support adhesion of various cell types assessed.

Modulation of prostate cancer cell attachment to matrix by versican.

In this study, we examined whether versican, a recognized anti-cell adhesive molecule for various mesenchymal and nerve cell types, influences prostate cancer cell adhesion to extracellular matrix components. Prostate cancer cell adhesion to fibronectin, a major component of the stromal extracellular matrix was inhibited by versican-rich conditioned medium (CM) from cultured human prostatic fibroblasts. In contrast, cancer cell attachment to laminin, a component of basement membranes, was not affected by the same CM.

The extracellular matrix protein betaIG-H3 is expressed at myotendinous junctions and supports muscle cell adhesion.

Molecules of the extracellular matrix (ECM) play important roles in the development and maintenance of myotendinous junctions (MTJs), specialized regions of muscle to bone union. In this report we provide evidence that skeletal muscle cells synthesize the collagen- and fibronectin-binding ECM protein betaIG-H3 and that betaIG-H3 is localized to MTJs. In situ hybridization experiments revealed that during E16.

An integrin binding peptide reduces rat CA1 hippocampal long-term potentiation during the first few minutes following theta burst stimulation.

The integrin binding peptide GRGDSP was tested on Schaffer to CA1 (Sch-CA1) long-term potentiation (LTP) in the rat hippocampal slice. Experiments in which GRGDSP was bath applied for 50 min, beginning 10 min before theta burst stimulation (TBS), reduced LTP of the field excitatory post synaptic potential in a concentration dependent manner, with 250 microM producing a significant decrease. However, LTP was not affected when 250 microM GRGDSP was applied 30 min post-TBS, nor when applied as soon as 5 min post-TBS.

Temporal effects of cell adhesion on mechanical characteristics of the single chondrocyte.

Cell adhesion to material surfaces is a fundamental phenomenon in tissue response to implanted devices, and an important consideration in tissue engineering. For example, elucidation of phenomena associated with adhesion of chondrocytes to biomaterials is critical in addressing the difficult problem of articular cartilage regeneration. The first objective of this study was to measure the mechanical adhesiveness characteristics of individual rabbit articular chondrocytes as a function of seeding time to provide further understanding of the cell adhesion process.

Quantification of varying adhesion levels in chondrocytes using the cytodetacher.

The potential to promote cell adhesion must be evaluated in the development of tissue-engineered implants and scaffolds. One measure of cell adherence is the force necessary to detach the cell. The objective of this study was to evaluate the cytodetacher (Athanasiou, K.

Regulation of stromal versican expression by breast cancer cells and importance to relapse-free survival in patients with node-negative primary breast cancer.

Purpose: Determination of meaningful prognostic indicesremains a high priority for women diagnosed with node-negative primary breast cancer. Currently, 30% of these women relapse, and there is no reliable means of predicting this group of patients. This study investigates whether the level of expression of versican, an anticell adhesive proteoglycan, in the peritumoral stromal tissue of women with node-negative, primary breast cancer predicts relapse-free survival.