The effect of carbonylcyanide m-chlorophenylhydrazone (CCCP) on the performance of isolated retrograde-perfused rabbit hearts was investigated in the present study. CCCP was first investigated as the blocking agent of photosynthesis. CCCP is also commonly used as the metabolic inhibitor of energy production. The animals were heparinized and anesthetized, the hearts were quickly removed and perfused with Krebs-Henseleit buffer in Langendorff's mode. Two parallel perfusion systems were used to distinguish the control and the treated heart. Two different concentrations of CCCP, low-dose (0.1 μM) and high-dose (0.5 μM), were used to study the dose-dependent relationship. The hemodynamic parameters used in the present study are aortic pressure (AOP), end diastolic pressure (EDP), peak systolic pressure (PSP), left ventricular developed pressure (LVDP), the rate of change of left ventricular pressure (positive dP/dt & negative dP/dt), and heart rate (HR). In general, CCCP impaired the mechanical performance of the heart by decreasing cardiac contractility. Positive and negative dP/dt were decreased 49.4% and 55.6%, respectively, by the low-dose of CCCP. High-doses of CCCP also decreased positive dP/dt and negative dP/dt by 81.4% and 88.9%, respectively. In addition, low-dose CCCP caused decreases of peak systolic pressure and left ventricular pressure developed by 50.4% and 61.6%. Similarly, high-dose CCCP decreased PSP and LVDP by 74.8% and 92.5%, respectively. The end diastolic pressure was increased 66.8% by low-dose CCCP. CCCP had no significant effects on aortic pressure and the heart rate. In conclusion, CCCP impaired the mechanical performance of the isolated perfused hearts as evidenced by decreasing PSP, LVDP, positive dP/dt and negative dP/dt. This degradation of myocardial performance showed a dose-dependent relationship. CCCP also caused a higher incidence rate of arrhythmia. Because CCCP uncoupled the electron transport from the ATP production in the mitochondria, the present study suggested that the development of contracture and heart failure was due to the energy depletion by CCCP.

Until the past few years, cellulose derivatives, (e.g., cellophane, collodion, and parchment) and animal membranes (e.g., goldbeater’s skin) have been the only dialysis membranes employed commercially. Cellophane has been used as the dialysis membrane in the artificial kidney since the machine’s inception in 1914. It continues to serve in this capacity, even though, in the last few years, attempts have been made to develop better films. An appreciation of both the “solution theory” and the “pore theory” is in order for this discussion. By incorporating the dynamics of these theories in the techniques of membram formulation, improved membrane performance can be exhibited. In the following discussion which considers membrane formulation, improved membrane performance can be displayed by higher particle transfer rates and greater particle selectivity.

In recent years there has been a growing emphasis on the development of health education in the public schools. This emphasis has been due largely to the realization of the importance and need for training in healthful living. As the educational system in the United States has assumed more and more responsibility for the total development of the child, physically, mentally, specially, and emotionally, educators and parents have felt the need for a more functional health program in the school.

The bZIP transcription factors make up a family of long α-helical proteins that dimerize based on a pattern of hydrophobic residues and bind to DNA through a region of basic residues. Because binding specificity is a particular topic of interest, the dimerization interaction is attractive as a possible candidate to better understand protein quaternary structure. Use of the Knob-Socket (KS) model for determination of packing structure provides a novel approach to analyze protein-protein interactions. A KS analysis of the protein-protein interface provides unique insight into the specificity of the classical leucine zipper pseudo-7mer repeat. From an analysis of the KS packing maps, this research provides evidence of a general framework for defining the specificity between coiled-coils. The KS maps show how hydrophobic specificity is defined in the coiled-coil interface, where knobs are centralized in the middle of the socket packing, while the peripheral socket residues are hydrophilic. Based on this KS analysis, the KS model will be used to design proteins that mimic the leucine zipper region of bZIP proteins. The proteins will be purified into E. coli and its 2º structure will be confirmed through circular dichroism. Binding specificity will be studied through mutations of the designed proteins and compared using the BACTH (bacterial adenylate cyclase two-hybrid) system.

Peptides have been used as targeting ligands in targeted drug delivery. Conjugating peptides to cytotoxicity agents via a linker to build peptide-drug conjugate (PDC) is a promising targeting strategy. The binding affinity of the peptide ligand and the receptor plays a crucial role in the PDC targeted drug delivery. Although the ligand binding which can be used in targeted drug delivery has been established conceptually, the quantitative or semi-quantitative contribution of binding affinity in targeting efficiency has not been fully explored. The optimal range of binding affinity of the peptide for targeted delivery remains unknown. Therefore, there is a lack of knowledge on the relationship between the peptide binding affinity and targeted drug delivery efficiency. The major steps in peptide drug delivery include cellular binding, cellular internalization, and tumor cells killing. In this study, three EGFR-targeting peptides with binding affinity levels ranging from 22 nM to 1.25 μM were selected to study their targeted drug delivery efficiency. The cellular binding study of FITC labeled peptides showed that peptide GE11 with the highest binding affinity had the highest cellular binding among three peptides. PEP11 peptide showed enhanced cellular binding compared to the L1 peptide. Moreover, GE11 also showed the selectivity of cellular binding between EGFR-positive cells and EGFR-negative cells. The cellular distribution showed that GE11-FITC could be successfully internalized into cells. The uptake mechanism studies demonstrated that the cellular uptake of GE11-FITC was based on receptor-mediated endocytosis, meaning that the cellular binding of GE11 was able to trigger the endocytosis. MMAE, a non-selective anticancer agent, was conjugated to the peptides through a protease-sensitive linker. The cytotoxicity assay showed that GE11-MMAE had the highest drug delivery efficiency and selectivity of three peptides, with 200 folds lower IC50 value than MMAE in EGFR-positive cells and 1000 times lower in EGFR-negative cells. PEP11-MMAE also showed an enhanced drug delivery than MMAE and L1-MMAE. L1-MMAE failed to show a significant difference with MMAE. Cellular binding kinetics results revealed that GE11-FITC had a higher rate of cellular uptake than PEP11-FITC. In conclusion, in the range from micromolar to the nanomolar, higher binding affinity of peptide ligand will contribute to higher cellular binding, targeted drug delivery efficiency, and cellular uptake rate. These results suggest that in EGFR-targeting delivery, the nanomolar level binding affinity is necessary for peptides to be used as targeting moiety in the targeted drug delivery. This study provides a starting point for further quantitative probing of the optimal binding affinity for designing and developing peptide ligand-based targeted delivery.

The end of the human chromosome is protected by telomeres which contain a special tandem guanine-rich DNA sequence, 5’-TTAGGG. The length of telomeres is shortened during cell replications, and its length limits the replication capacity of cells. Telomerase is over-expressed in 85–90% of cancer cells, responsible for extending the telomere length in cancer cells. Guanine-rich DNA sequence can self-assemble into unique G-quadruplex structures that interfere with the extension of telomeres by telomerase. Therefore, DNA G-quadruplex has recently received much attention because of its important regulatory functions in telomerase-mediated cancerization. The formation of G-quadruplex requires monovalent cations (Na+ and K+) or small molecules known as G-quadruplex ligands. In the present work, we developed a serial of G-quadruplex ligands by tethering side-chains to two core structures: 1,10-phenanthroline (Phen) and thiazole orange (TO). Biophysical studies including DNA thermal denaturation monitored by fluorescence orcircular dichroism, fluorometric titration, and ESI-MS spectrometry reveal that the binding of the synthesized ligands to G-quadruplex is side-chain dependent. The arylsulfanyl side chains significantly improve the binding affinity and selectivity of 1,10-phenanthroline towards G-quadruplex over duplex DNA. The polyamine side chains are a suitable structural motif for remarkable G-quadruplex binding affinity based on the results from both Phen and TO derivatives. These ligands greatly inhibit the telomerase activity in vitro, determined by a modified telomeric repeat amplification protocol (TRAP) assay. Amongst these promising telomerase inhibitors, a thiazole orange derivative containing a side chain of spermine shows an outstanding telomerase inhibition effect at nanomolar concentrations, which is comparable to the most effective synthetic telomerase inhibitors, BRACO-19.

The intent of this thesis is to explore the reasons why young athletes may conceal their sports injuries. In recent years, there has been much discussion about the long-term health implications that former athletes are dealing with as they live life after sports. Sports injuries including concussions, knee damage, and spinal injuries are all issues that could affect an athlete’s quality of life far beyond their playing days. It is well known around the athletic and medical communities that many athletes withhold information about their injury symptoms just to get back on to the field. Most worrisome about this fact, is the disregard of any long-term damage being done to their body. In this study, we explored the influence of social norms, perceived masculinity, and other external influences on athlete populations in an attempt to understand the reasons why injuries are so often under-reported and masked by athletes. Understanding the logic behind why athletes “play through” injuries and the external influences that may cause this behavior, is essential to athlete safety in the future. Results showed significant findings among highly masculine athletes and injury concealment as well as in athletes who feared losing a performance role and injury concealment. Those athletes who wish to appear tough and masculine as well as those athletes who may be fearful of losing a performance role were more likely to conceal their injuries. Athletes who are a part of a team were also likely to behave in the same way and understanding these reasons can help improve athlete safety in the years to come.

Pancreatic cancer is the product of microenvironment alterations which emerge from inflammatory signaling and progress to more devastating cases such as Pancreatic Ductal Adenocarcinoma (PDAC). PDAC is extremely aggressive with a statistical five-year survival rate of merely 3%-5%, and is more than relevant to cancer research being that it is the fourth leading cause of cancer-related deaths in the US. Unfortunately pancreatic cancer is often unnoticed until reaching its hardly treatable end stages, which perpetuates the low survival rate. The onset of PDAC may be facilitated by the activation of pancreatic stellate cells (PSCs), which secrete collagen and markedly contribute to tissue fibrosis. Inflammatory factors and activation of PSCs are hallmarks of pancreatitis and could increase occurrence rates of pancreatic cancer. The purpose of this thesis is to elucidate inflammatory signaling patterns starting with the onset of acute pancreatitis and through future studies of the more damaging states of chronic pancreatitis and cancer progression. Through the induction of acute pancreatitis in oncogenic and wild type mouse models and evaluating cytokine expression levels via RT-PCR a link between inflammatory signaling and disease state progression will be delineated. This model utilizes mice with mutant KRas, a gene activated in nearly all PDAC incidences, and constitutively active Akt, an oncogene activated in nearly all cancers. Preliminary results indicate that when experimentally inducing pancreatitis in mice predisposed to pancreatic cancer tissue remodeling and leukocyte infiltration is observed as a result of cytokine expression. Furthermore, macrophage and neutrophil stains are positive with one round of cerulein injections proving that acute inflammation is induced by these methods. Pancreatitis is a risk factor for pancreatic cancer which can be caused by environmental factors including smoking, alcohol consumption, and obesity. By understanding the mechanism by which inflammation occurs and the cytokine signaling involved we can attempt inhibit tumor-promoting signaling pathways in the pancreas stroma.

The treatment of most cancers can still be considered inadequate despite the steady progress being made. A prime example of this issue is with epithelial ovarian cancers; this disease presents a significant issue, with a 5-year survival rate of 46% and a survival rate of 28% in patients that develop metastatic disease. Since ovarian cancer has such a high mortality rate, effective treatment modalities are necessary to prolong the quality of life after diagnosis. Psychosocial stress is related to the progression, proliferation, and migration in cancer patients, but the mechanisms of this relationship are not fully understood. The present in vitro study investigated the ability of oxytocin, a neuropeptide associated with social support, to attenuate the stress response. Catecholamines, a subclass of stress hormones, were used to simulate the stress induced inflammation process in ovarian cancer cells. To evaluate oxytocin’s capacity to attenuate the stress response, the ovarian cancer cell lines SKOV3, HEYA8, OVCAR8, and OV432 were separately treated with the presence or absence of catecholamines with the addition of oxytocin. Protein expression of the oxytocin receptor was investigated using a western blot protocol. Oxytocin receptor, oxytocin, and IL-6 mRNA expression was evaluated by quantitative PCR. Treatment with Oxytocin attenuated the inflammatory response resulting from catecholamine treatment. The oxytocin receptor gene and protein were present in each cell line, suggesting that oxytocin has an anti-inflammatory role in the tumor microenvironment in ovarian cancer patients. These results provide a mechanism by which social support, working through the release of oxytocin, promotes an anti-inflammatory process in ovarian cancer patients. This study may shed light into new pharmacological approaches for the treatment of ovarian cancer.

PURPOSE: The primary aim of this study was to examine the accuracy of a hip (Evenson algorithm) and wrist-worn (Crouter algorithm) accelerometer in assessing time spent in different intensity categories in pre-adolescent girls during semi-structured dance classes using direct observation (D.O.) as the criterion measure. The secondary aim of this study was to examine the validity of a wrist-worn accelerometer for dichotomizing pre-adolescent girls as meeting or not meeting different preselected doses of moderate-to-vigorous PA compared to the hip-worn accelerometer. METHODS: Data were collected and analyzed on a total of 6 participants (age = 10.22 ± 2.38) for the primary aim. Additionally, data was collected and analyzed on a total of 20 participants (age = 8.6 ± 1.6) for the secondary aim. RESULTS: Compared to D.O., the wrist-worn accelerometer was inaccurate in measuring time spent in light PA, vigorous PA and MVPA. Additionally, the hip-worn accelerometer was inaccurate in measuring time spent in sedentary time, light PA, vigorous PA and total PA. Further, for the secondary aim, there was a significant difference between device location and meeting PA dosage for three days and five days. CONCLUSION: Traditional accelerometer algorithms rely on the activity count cut-point method which provides mixed to poor results of activity intensity classification regardless of wear location. Future research should move away from the activity count cut-point method and aim to develop algorithms that use more of the rich data available from the accelerometers’ acceleration signal.