No description available.

This thesis describes the synthesis of a novel multimeric peptide ligand targeted to the human melanocortin 4 receptor. The synthesis of the peptide was attempted both by solid phase peptide synthesis and by solution phase peptide synthesis, leading to the conclusion that the necessary C- and N- terminal substituents were much easier to install via the solution phase route. The bifunctional peptide was purified and then multimerized in both protected and active amino acid forms using the copper(I)-catalyzed azide alkyne cycloaddition (CuAAC) reaction. The multimers were characterized using MS and UV-Vis spectroscopy. It was found that a large portion of the monomer cyclized under CuAAC conditions, though sufficient multimerization took place to form up to nonamers, as determined by mass spectrometry.

Human tumor cell progression and metastasis is partially dependent on the ability of tumor cells to adhere to the proteins of the extracellular matrix and migrate to distant locations. Using a combinatorial screening approach, six novel D-amino acid containing peptides were identified and analyzed for their ability to adhere to human prostate tumor cells, support tumor cell adhesion and inhibit tumor cell adhesion to ECM proteins. Two peptides, RZ-3 (kmviywkag) and HYD1 (kikmviswkg) bound to tumor cell surfaces. A scrambled peptide derivative of HYD1, HYDS (wiksmkivkg) is not active. As immobilized ligands, RZ-3 and HYD1 can support prostate tumor cell adhesion. Prostate tumor cell adhesion to immobilized RZ-3 and HYD1 is integrin dependent. Soluble RZ-3 and HYD1 inhibits tumor cell adhesion to extracellular matrix proteins in a concentration dependent manner. These results indicate that RZ-3 and HYD1 are biologically active D-amino acid containing peptides that can support tumor cell adhesion and can inhibit tumor cell adhesion to immobilized extracellular matrix proteins.Cell migration is dependent on adhesive interactions with the extracelluar matrix. These interactions induce signaling and cytoskeletal responses necessary for migration. HYD1 completely blocks random haptotactic migration and inhibits invasion of prostate carcinoma cells on laminin-5. This effect is adhesion independent and reversible. The inhibition of migration by HYD1 involves a dramatic remodeling of the actin cytoskeleton resulting in increased stress fiber formation and actin colocalization with cortactin at the cell membrane. HYD1 interacts with a6 and a3 integrin subunits and elevates laminin-5 dependent intracellular signals including focal adhesion kinase, mitogen activated protein kinase kinase, and extracellular signal-regulated kinase. The scrambled derivative of HYD1, HYDS, does not interact with the a6 or a3 integrin subunits and is not biologically active. The minimal element for bioactivity of HYD1 was determined using alanine-substituted analogs of HYD1 and N- and C-terminal deletion mutants of HYD1. The minimal element necessary to block cell migration on laminin-5 and activate cell signaling through ERK is xikmviswxx. Taken together, these results indicate that HYD1 is a biologically active integrin-targeting peptide that reversibly inhibits tumor cell migration on laminin-5 and uncouples phosphotyrosine signaling from cytoskeletal dependent migration.

The anterior cingulate cortex (ACC) expresses high density of Gastrin-releasing peptide (GRP) and GRP receptor mRNA. To address possible function, this investigation used patch clamp recordings in mouse brain slice preparations to evaluate intrinsic properties of ACC neurons and neuronal responses to bath-applied GRP peptide. The ACC neurons were divided according to their morphology, the properties of action potentials and their firing pattern in response to depolarizing current pulses. Two physiological groups of interneurons and three groups of pyramidal neurons were defined. Application of the GRP induced depolarization and increased firing of the interneurons while hyperpolarization and reduced firing in pyramidal neurons. Moreover, activation of GRP receptor facilitated GABAergic neurotransmission via a postsynaptic mechanism. The results suggest that GRP receptor is an important regulator of neuronal circuits in the ACC and may consequently play an important role for ACC neurons in the central processing of high brain function.

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No description available.

The natriuretic peptide system (NPS) is a hormonal system critical to mammalian cardiovascular homeostasis. The purpose of the present study was to investigate the role of ANP during early postnatal cardiac development by i) monitoring the development of cardiac hypertrophy during early postnatal development of the ANP-/- mice, and ii) comparing morphologic, morphometric and molecular differences in ANP+/+ mice compared to ANP-/- mice during this developmental period. Age matched male ANP+/+ and ANP-/- mice, aged day 1 and weeks 1 to 5, were evaluated. Body weight, organ weights and hematocrit were recorded. RNA was isolated and quantitative real-time RT-PCR was used to monitor cardiac gene expression. An additional cohort of animals was used for morphologic and morphometric analysis. Heart weight to body weight ratio (HW/BW) was dramatically higher in ANP-/- animals at all time points, indicating cardiac hypertrophy is established before the advent of adult blood pressure. Molecular analysis of gene expression revealed a compensatory response of the NPS in the ANP-/- mice. Specifically an up-regulation of BNP expression in ANP-/- mice was noted throughout postnatal development. Similarly, NPR-A and NPR-C demonstrated compensatory action for the lack of ANP, as expressional levels also varied throughout development. Morphological analysis of cardiac vasculature revealed striking structural differences between ANP+/+ and ANP-/- mice. Quantitative stereological analysis of LM images indicated a greater vessel volume in ANP-/- compared to ANP+/+ mice. This study demonstrates that alterations in early molecular events, such as changes in NPS expression, may be responsible for the maintenance and progression of cardiac hypertrophy during early postnatal development in the ANP-/- mice. The absence of ANP during this critical period of development has a profound impact on final cardiac structure leading to future pathological states. / Thesis (Master, Anatomy & Cell Biology) -- Queen's University, 2010-02-05 14:15:33.982

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Amylin is a 37 amino acid long peptide that aggregates into toxic oligomers and fibrils. Since amylin is secreted by and also acts on pancreatic beta cells, type II diabetes is classified as an amyloidogenic disease. This study focuses on the development of a cost effective chemical synthetic strategy for amylin synthesis as previous studies relied on extremely expensive pseudoproline derivatives. Furthermore, commercially available amylin varies between batches and also contains impurities that could generate anomalies and affect reproducibility of experiments. Secondly, chemically synthesized non-methylated and N-methylated derivatives of amylin were shown to inhibit toxicity of full length amylin. A fluorescentlylabeled chemically synthesized derivative of amylin was used to track cellular localization of amylin via confocal microscopy. Amylin aggregation kinetics was established using a surface plasmon resonance (SPR) biosensor. In addition, nanoparticle tracking analysis (NTA) was used as a novel technique to determine the size of oligomers over real time. This technology indicated that the size range of the toxic species of amylin is between 200-300 nm. Furthermore, it can be suggested that NTA could potentially be developed into a screening tool for inhibitors of amylin-mediated cytotoxicity. / Thesis (Ph.D.)-University of KwaZulu-Natal, Westville, 2012.

The anterior cingulate cortex (ACC) expresses high density of Gastrin-releasing peptide (GRP) and GRP receptor mRNA. To address possible function, this investigation used patch clamp recordings in mouse brain slice preparations to evaluate intrinsic properties of ACC neurons and neuronal responses to bath-applied GRP peptide. The ACC neurons were divided according to their morphology, the properties of action potentials and their firing pattern in response to depolarizing current pulses. Two physiological groups of interneurons and three groups of pyramidal neurons were defined. Application of the GRP induced depolarization and increased firing of the interneurons while hyperpolarization and reduced firing in pyramidal neurons. Moreover, activation of GRP receptor facilitated GABAergic neurotransmission via a postsynaptic mechanism. The results suggest that GRP receptor is an important regulator of neuronal circuits in the ACC and may consequently play an important role for ACC neurons in the central processing of high brain function.