Development of an injectable slow release system for bone morphogenetic protein-2

In this thesis, development of a bone regeneration therapy using biomaterials and growth factors is described. Initially, collagen I based recombinant protein (RCP) microspheres were developed for BMP-2 delivery. Among several parameters investigated, size and crosslinking of microspheres affected the BMP-2 release. To improve bone formation, we have combined microspheres with in-situ gelling hydrogels. Two types of alginate, high mannuronate (SLM) and high guluronate (SLG), and one type of thermosensitive hyaluronic acid were developed for this study. These hydrogels were designed not only to deliver and keep the microspheres at the site but also to act as a scaffold for the regenerating tissue and to fill a defect. The SLG alginate-RCP microspheres formulation was selected for further studies in ectopic and orthotopic bone formation models in rats. First, the effect of BMP-2 dose delivered was tested in the ectopic bone model using 10, 3, 1, 0.3 and 0 μg BMP-2 per implant. In a subsequent calvarial defect experiment, 50 μg/mL and 5 μg/mL BMP-2 containing hydrogels were tested. This study showed that the effective concentration of BMP-2 to induce both ectopic and orthotopic bone is optimally between 15-50 μg/mL

Driving behavior classification for Heavy-Duty vehicles using LSTM Networks

Despite growing autonomous driving trend, human is still a major factor in the current vehicle technology. Drivers have a great impact on both fuel economy and accident prevention. Therefore, identi cation and evaluation of driving behaviors are crucial to improve the performance, safety and energy management of vehicle technologies, particularly for heavy-duty vehicles. In this thesis, several driving behaviors with di erent acceleration and car following characteristics are generated on a realistic truck model in IPG's TruckMaker simulation environment. A Long Short Term Memory (LSTM) classi er is then utilized to recognize driving behaviors. First, six drivers are de ned based on their longitudinal and lateral acceleration limits. The classi er is trained using driving signals acquired from the simulated truck which follows an arti cial training road with di erent trailer loads. The training road is designed to cover possible road curves that can be seen in highways. The model is tested with driving signals that are collected from a realistic road using the same method. Then, three drivers (calm, normal and aggressive) are de ned based on their longitudinal acceleration pro les in car following and the classi er is trained and tested using driving signals of these drivers in di erent tra c scenarios. Results show that the proposed LSTM classi er is capable of successfully capturing the dynamic relations encoded in driving signals and recognizing di erent driving behaviors in small time sample

Quantitative Sequencing for the Determination of Kdr-type Resistance Allele (V419L,L925I, I936F) Frequencies in Common Bed Bug, Cimex lectularius L., (Hemiptera: Cimicidae) Populations Collected from Israel

Human bed bug infestations have dramatically increased worldwide since the mid-1990s. A similar phenomenon was also observed in Israel since 2005, when infestations were reported from all over the country. Two single nucleotide polymorphisms (V419L and L925I) in the bed bug voltage sensitive sodium channel confer kdr-type resistance to pyrethroids. Using quantitative sequencing (QS), the resistance allele frequencies of Israeli bed bug populations from across the country were determined. Genomic DNA was extracted from samples of 12 populations of bed bugs collected from Israel and DNA fragments containing the V419L or L925I and I936F mutations sites were PCR amplified. The PCR products were analyzed by QS and the nucleotide signal ratios calculated and used to predict the resistance allele frequencies of the unknown populations. Results of the genetic analysis show that resistant nucleotide signals are highly correlated to resistance allele frequencies for both mutations. Ten of the 12 tested populations had 100% of the L925I mutation and 0% of the V419L mutation. One population was heterogeneous for the L925I mutation and had 0% of the V419L mutation and another population was heterozygous for the V419L mutation and had 100% of the L925I mutation. I936F occurred only at low levels. These results indicate that bed bugs in Israel are genetically resistant to pyrethroids. Thus, pyrethroids should only be used for bed bug management with caution using effective application and careful monitoring procedures. Additionally, new and novel-acting insecticides and non-chemical means of controlling bed bugs should be explored

Oligonucleotide delivery with cell surface binding and cell penetrating peptide amphiphile nanospheres

Cataloged from PDF version of article.A drug delivery system designed specifically for oligonucleotide therapeutics can ameliorate the problems associated with the in vivo delivery of these molecules. The internalization of free oligonudeotides is challenging, and cytotoxicity is the main obstacle for current transfection vehicles. To develop nontoxic delivery vehicles for efficient transfection of oligonudeotides, we designed a self-assembling peptide amphiphile (PA) nanosphere delivery system decorated with cell penetrating peptides (CPPs) containing multiple arginine residues (R-4 and R-8), and a cell surface binding peptide (KRSR), and report the efficiency of this system in delivering G-3129, a Bcl-2 antisense oligonucleotide (AON). PA/AON (peptide amphiphile/antisense oligonucleotide) complexes were characterized with regards to their size and secondary structure, and their cellular internalization efficiencies were evaluated. The effect of the number of arginine residues on the cellular internalization was investigated by both flow cytometry and confocal imaging, and the results revealed that uptake efficiency improved as the number of arginines in the sequence increased. The combined effect of cell penetration and surface binding property on the cellular internalization and its uptake mechanism was also evaluated by mixing R-8-PA and KRSR-PA. R-8 and R-8/KRSR decorated PAs were found to drastically increase the internalization of AONs compared to nonbioactive PA control. Overall, the KRSR-decorated self-assembled PA nanospheres were demonstrated to be noncytotoxic delivery vectors with high transfection rates and may serve as a promising delivery system for AONs