Animal Experimentation and Research Commons^™

Optimization Of Primary Cilia Detection In Musculoskeletal Tissue, Ragen Engel

Williams Honors College, Honors Research Projects

The objective of this research project was to optimize a laboratory technique for visualizing primary cilia in musculoskeletal tissue of young swine, specifically the growth plate of long bones. This was accomplished through varying fixation procedures, antigen retrieval methods, antibody concentrations, and incubation times. By varying these parameters, a reproducible procedure was developed to examine the primary cilia in multiple tissues including ligament, fascia, and growth plate. This experiment introduced variances in fixation methods, with methanol or formalin. Paraffin and frozen embedding techniques were also varied for comparison with regards to cilia visualization. Results from the acetylated alpha tubulin (aTUB) …

Development Of A Pd-L1 Pet Imaging Biomarker, Caleb Jack Bridgwater

Posters-at-the-Capitol

Immunotherapy strategies are very promising treatments for cancer patients. Specifically, Immune checkpoint inhibitor therapy focusing on the PD-1/PD-L1 pathway shows long-lasting positive results in many cancer patients. Unfortunately, not all the patients can benefit from this highly effective treatment. Hence, there is a great need for predictive biomarkers. Immunohistochemical (IHC) staining has been used as a way of predicting patient response, yet shows many problems. For example, IHC utilizes an invasive biopsy and sample fixing, which creates an incomplete and delayed picture of the patient’s biochemistry and the tumor microenvironment, consequently ignoring metastases.

The purpose of this study is to …

Structural Insights Into The Potency Of Sk Channel Positive Modulators, Young-Woo Nam, Razan Orfali, Tingting Liu, Kunqian Yu, Meng Cui, Heike Wulff, Miao Zhang

Pharmacy Faculty Articles and Research

Small-conductance Ca2+-activated K+ (SK) channels play essential roles in the regulation of cellular excitability and have been implicated in neurological and cardiovascular diseases through both animal model studies and human genetic association studies. Over the past two decades, positive modulators of SK channels such as NS309 and 1-EBIO have been developed. Our previous structural studies have identified the binding pocket of 1-EBIO and NS309 that is located at the interface between the channel and calmodulin. In this study, we took advantage of four compounds with potencies varying over three orders of magnitude, including 1-EBIO, NS309, SKS-11 (6-bromo-5-methyl-1H-indole-2,3-dione-3-oxime) and …

Theta-Burst Microstimulation In The Human Entorhinal Area Improves Memory Specificity, Ali S. Titiz, Michael R. H. Hill, Emily A. Mankin, Zahra M. Aghajan, Dawn Eliashiv, Natalia Tchemodanov, Uri Maoz, John Stern, Michelle E. Tran, Peter Schuette, Eric Behnke, Nanthia A. Suthana, Itzhak Fried

Psychology Faculty Articles and Research

The hippocampus is critical for episodic memory, and synaptic changes induced by long-term potentiation (LTP) are thought to underlie memory formation. In rodents, hippocampal LTP may be induced through electrical stimulation of the perforant path. To test whether similar techniques could improve episodic memory in humans, we implemented a microstimulation technique that allowed delivery of low-current electrical stimulation via 100 μm-diameter microelectrodes. As thirteen neurosurgical patients performed a person recognition task, microstimulation was applied in a theta-burst pattern, shown to optimally induce LTP. Microstimulation in the right entorhinal area during learning significantly improved subsequent memory specificity for novel portraits; participants …

Efficacy And Safety Comparison Between Suberoylanilide Hydroxamic Acid And Mitomycin C In Reducing The Risk Of Corneal Haze After Prk Treatment In Vivo, Govindaraj Anumanthan, Ajay Sharma, Michael R. Waggoner, Chuck W. Hamm, Suneel Gupta, Nathan P. Hesemann, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

PURPOSE:

This study compared the efficacy and safety of suberoylanilide hydroxamic acid (SAHA) and mitomycin C (MMC) up to 4 months in the prevention of corneal haze induced by photorefractive keratectomy (PRK) in rabbits in vivo.

METHODS:

Corneal haze in rabbits was produced with −9.00 diopter PRK. A single application of SAHA (25 μM) or MMC (0.02%) was applied topically immediately after PRK. Effects of the two drugs were analyzed by slit-lamp microscope, specular microscope, TUNEL assay, and immunofluorescence.

RESULTS:

Single topical adjunct use of SAHA (25 μM) or MMC (0.02%) after PRK attenuated more than 95% corneal …

Mammary Extracellular Matrix Directs Differentiation Of Testicular And Embryonic Stem Cells To Form Functional Mammary Glands In Vivo, Robert D. Bruno, Jodie M. Fleming, Andrea L. George, Corinne A. Boulanger, Pepper Schedin, Gilbert H. Smith

School of Medical Diagnostics & Translational Sciences Publications

Previously, we demonstrated the ability of the normal mammary microenvironment (niche) to direct non-mammary cells including testicular and embryonic stem cells (ESCs) to adopt a mammary epithelial cell (MEC) fate. These studies relied upon the interaction of transplanted normal MECs with non-mammary cells within the mammary fat-pads of recipient mice that had their endogenous epithelium removed. Here, we tested whether acellular mammary extracellular matrix (mECM) preparations are sufficient to direct differentiation of testicular-derived cells and ESCs to form functional mammary epithelial trees in vivo. We found that mECMs isolated from adult mice and rats were sufficient to redirect testicular derived …

Epigenetic Modification Prevents Excessive Wound Healing And Scar Formation After Glaucoma Filtration Surgery, Ajay Sharma, Govindaraj Anumanthan, Marcos Reyes, Huiyi Chen, Jason W. Brubaker, Saad Siddiqui, Suneel Gupta, Frank G. Rieger, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

PURPOSE. The purpose of this study was to determine the efficacy of suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor (HDACi), in prevention of excessive wound healing and scar formation in a rabbit model of glaucoma filtration surgery (GFS).

METHODS. A rabbit model of GFS was used. Rabbits that underwent GFS received balanced salt solution, or SAHA (50 lM), or mitomycin C (0.02%). Clinical scores of IOP, bleb vascularity, and slit-lamp examination were performed. On postoperative day 14, rabbits were killed and the bleb tissues were collected for evaluation of tissue fibrosis with hematoxylin and eosin, Masson trichrome, a-smooth muscle …

Development Of A Novel In Vivo Corneal Fibrosis Model In The Dog, K. M. Gronkiewicz, Elizabeth A. Giuliano, K. Kuroki, F. Bunyak, Ajay Sharma, L. B. C. Teixeira, C. W. Hamm, R. R. Mohan

Pharmacy Faculty Articles and Research

The aim of this study was to develop a novel in vivo corneal model of fibrosis in dogs utilizing alkali burn and determine the ability of suberanilohydroxamic acid (SAHA) to inhibit corneal fibrosis using this large animal model. To accomplish this, we used seven research Beagle dogs. An axial corneal alkali burn in dogs was created using 1 N NaOH topically. Six dogs were randomly and equally assigned into 2 groups: A) vehicle (DMSO, 2 μL/mL); B) anti-fibrotic treatment (50 μM SAHA). The degree of corneal opacity, ocular health, and anti-fibrotic effects of SAHA were determined utilizing the Fantes grading …

Molecular Mechanisms Of Suberoylanilide Hydroxamic Acid In The Inhibition Of Tgf-Β1-Mediated Canine Corneal Fibrosis, Kristina M. Gronkiewicz, Elizabeth A. Giuliano, Ajay Sharma, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

Objective—To investigate molecular mechanisms mediating anti-fibrotic effect of SAHA in the canine cornea using an in vitro model. We hypothesized that SAHA attenuates corneal fibrosis by modulating Smad-dependent and, to a lesser extent, Smad-independent signaling pathways activated by TGF-β1, as well as matrix metalloproteinase (MMP) activity.

Methods—Cultured canine corneal fibroblasts (CCF) were incubated in the presence/absence of TGF-β1 (5ng/ml) and SAHA (2.5μM) for 24hrs. Western blot analysis was used to quantify non-phosphorylated and phosphorylated isoforms of Smad2/3, p38 MAP kinase (MAPK), ERK1/2 and JNK1. Real-time PCR and zymography were utilized to quantify MMP1, MMP2, MMP8 and MMP9 mRNA expression and …

Role Of 5'Tg3'-Interacting Factors (Tgifs) In Vorinostat (Hdac Inhibitor)-Mediated Corneal Fibrosis Inhibition, Ajay Sharma, Nishant R. Sinha, Saad Siddiqui, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

Purpose: We have previously reported that vorinostat, an FDA-approved, clinically used histone deacetylase (HDAC) inhibitor, attenuates corneal fibrosis in vivo in rabbits by blocking transforming growth factor β (TGFβ). The 5′TG3′-interacting factors (TGIFs) are transcriptional repressors of TGFβ1 signaling via the Smad pathway. The present study was designed to explore the expression of TGIFs in human corneal fibroblasts and to investigate their role in mediating the antifibrotic effect of vorinostat.

Methods: Human corneal fibroblast cultures were generated from donor corneas. RNA isolation, cDNA preparation, and PCR were performed to detect the presence of TGIF1 and TGIF2 transcripts. The cultures were …

Paracrine-Rescued Lobulogenesis In Chimeric Outgrowths Comprising Progesterone-Receptor-Null Mammary Epithelium And Redirected Wild-Type Testicular Cells, Robert D. Bruno, Corinne A. Boulanger, Sonia M. Rosenfield, Lisa H. Anderson, John P. Lydon, Gilbert H. Smith

School of Medical Diagnostics & Translational Sciences Publications

We have previously shown that non-mammary and tumorigenic cells can respond to the signals of the mammary niche and alter their cell fate to that of mammary epithelial progenitor cells. Here we tested the hypothesis that paracrine signals from mammary epithelial cells expressing progesterone receptor (PR) are dispensable for redirection of testicular cells, and that re-directed wild-type testicular-derived mammary cells can rescue lobulogenesis of PR-null mammary epithelium by paracrine signaling during pregnancy. We injected PR-null epithelial cells mixed with testicular cells from wild-type adult male mice into cleared fat-pads of recipient mice. The testicular cells were redirected in vivo to …

Bmp7 Gene Transfer Via Gold Nanoparticles Into Stroma Inhibits Corneal Fibrosis In Vivo, Ashish Tandon, Ajay Sharma, Jason T. Rodier, Alexander M. Klibanov, Frank G. Rieger, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

This study examined the effects of BMP7 gene transfer on corneal wound healing and fibrosis inhibition in vivo using a rabbit model. Corneal haze in rabbits was produced with the excimer laser performing -9 diopters photorefractive keratectomy. BMP7 gene was introduced into rabbit keratocytes by polyethylimine-conjugated gold nanoparticles (PEI2- GNPs) transfection solution single 5-minute topical application on the eye. Corneal haze and ocular health in live animals was gauged with stereo- and slit-lamp biomicroscopy. The levels of fibrosis [a-smooth muscle actin (aSMA), F-actin and fibronectin], immune reaction (CD11b and F4/80), keratocyte apoptosis (TUNEL), calcification (alizarin red, vonKossa and osteocalcin), and …

Late Developing Mammary Tumors And Hyperplasia Induced By A Low-Oncogenic Variant Of Mouse Mammary Tumor Virus (Mmtv) Express Genes Identical To Those Induced By Canonical Mmtv, Robert D. Bruno

School of Medical Diagnostics & Translational Sciences Publications

Background: The canonical milk-transmitted mouse mammary tumor virus (MMTV) of C3H mice (C3H-MMTV) rapidly induces tumors in 90% of infected animals by 8 months of age. Pro-viral insertions of C3H-MMTV into genomic DNA results in the overexpression of common core insertion site (CIS) genes, including Wnt1/10b, Rspo2, and Fgf3. Conversely, infection by either the endogenous Mtv-1 virus (in C3Hf) or the exogenous nodule-inducing virus (NIV) (in Balb/c NIV) induces premalignant mammary lesions and tumors with reduced incidence and longer latency than C3H-MMTV. Here, we asked whether Mtv-1/NIV affected the expression of core CIS genes.

Findings: We confirmed the presence of …

Vorinostat: A Potent Agent To Prevent And Treat Laser-Induced Corneal Haze, Ashish Tandon, Jonathan C. K. Tovey, Michael R. Waggoner, Ajay Sharma, John W. Cowden, Daniel J. Gibson, Yuanjing Liu, Gregory S. Schultz, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

PURPOSE—This study investigated the efficacy and safety of vorinostat, a deacetylase (HDAC) inhibitor, in the treatment of laser-induced corneal haze following photorefractive keratectomy (PRK) in rabbits in vivo and transforming growth factor beta 1 (TGFβ1) -induced corneal fibrosis in vitro.

METHODS—Corneal haze in rabbits was produced with −9.00 diopters (D) PRK. Fibrosis in cultured human and rabbit corneal fibroblasts was activated with TGFβ1. Vorinostat (25 μm) was topically applied once for 5 minutes on rabbit cornea immediately after PRK for in vivo studies. Vorinostat (0 to 25 μm) was given to human/rabbit corneal fibroblasts for 5 minutes or 48 …

Attenuation Of Corneal Myofibroblast Development Through Nanoparticle-Mediated Soluble Transforming Growth Factor-Β Type Ii Receptor (Stgfβrii) Gene Transfer, Ajay Sharma, Jason T. Rodier, Ashish Tandon, Alexander M. Klibanov, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

Purpose: To explore (i) the potential of polyethylenimine (PEI)-DNA nanoparticles as a vector for delivering genes into human corneal fibroblasts, and (ii) whether the nanoparticle-mediated soluble extracellular domain of the transforming growth factor–β type II receptor (sTGFβRII) gene therapy could be used to reduce myofibroblasts and fibrosis in the cornea using an in vitro model.

Methods: PEI-DNA nanoparticles were prepared at a nitrogen-to-phosphate ratio of 30 by mixing linear PEI and a plasmid encoding sTGFβRII conjugated to the fragment crystallizable (Fc) portion of human immunoglobulin. The PEI-DNA polyplex formation was confirmed through gel retardation assay. Human corneal fibroblasts (HCFs) were …

Targeted Decorin Gene Therapy Delivered With Adeno-Associated Virus Effectively Retards Corneal Neovascularization In Vivo, Rajiv R. Mohan, Jonathan C. K. Tovey, Ajay Sharma, Gregory S. Schultz, John W. Cowden, Ashish Tandon

Pharmacy Faculty Articles and Research

Decorin, small leucine-rich proteoglycan, has been shown to modulate angiogenesis in nonocular tissues. This study tested a hypothesis that tissue-selective targeted decorin gene therapy delivered to the rabbit stroma with adeno-associated virus serotype 5 (AAV5) impedes corneal neovascularization (CNV) in vivo without significant side effects. An established rabbit CNV model was used. Targeted decorin gene therapy in the rabbit stroma was delivered with a single topical AAV5 titer (100 μl; 5x10^12 vg/ml) application onto the stroma for two minutes after removing corneal epithelium. The levels of CNV were examined with stereomicroscopy, H&E staining, lectin, collagen type IV, CD31 immunocytochemistry and …

Polyethylenimine-Conjugated Gold Nanoparticles: Gene Transfer Potential And Low Toxicity In The Cornea, Ajay Sharma, Ashish Tandon, Jonathan C. K. Tovey, Rangan Gupta, J. David Robertson, Jennifer A. Fortune, Alexander M. Klibanov, John W. Cowden, Frank G. Rieger, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

This study examined the gene transfer efficiency and toxicity of 2-kDa polyethylenimine conjugated to gold nanoparticles (PEI2-GNP) in the human cornea in vitro and rabbit cornea in vivo. PEI2-GNP with nitrogen-to-phosphorus (N/P) ratios of up to 180 exhibited significant transgene delivery in the human cornea without altering the viability or phenotype of these cells. Similarly, PEI2-GNP applied to corneal tissues collected after 12 h, 72 h, or 7 days exhibited appreciable gold uptake throughout the rabbit stroma with gradual clearance of GNP over time. Transmission electron microscopy detected GNP in the keratocytes and the extracellular matrix of the rabbit corneas. …

Significant Inhibition Of Corneal Scarring In Vivo With Tissue-Selective, Targeted Aav5 Decorin Gene Therapy, Rajiv R. Mohan, Ashish Tandon, Ajay Sharma, John W. Cowden, Jonathan C. K. Tovey

Pharmacy Faculty Articles and Research

PURPOSE. This study tested a hypothesis that tissue-selective targeted decorin gene therapy delivered to the stroma with adeno-associated virus serotype 5 (AAV5) inhibits corneal fibrosis in vivo without significant side effects.

METHODS. An in vivo rabbit model of corneal fibrosis was used. Targeted decorin gene therapy was delivered to the rabbit cornea by a single topical application of AAV5 (100 L; 6.5 1012 g/mL) onto the bare stroma for 2 minutes. The levels of corneal fibrosis were determined with stereomicroscopy, slit lamp biomicroscopy, -smooth muscle actin ( SMA), fibronectin, and F-actin immunocytochemistry, and/or immunoblotting. CD11b, F4/80 immunocytochemistry, and TUNEL assay …

Efficacious And Safe Tissue-Selective Controlled Gene Therapy Approaches For The Cornea, Rajiv R. Mohan, Sunilima Sinha, Ashish Tandon, Rangan Gupta, Jonathan C. K. Tovey, Ajay Sharma

Pharmacy Faculty Articles and Research

Untargeted and uncontrolled gene delivery is a major cause of gene therapy failure. This study aimed to define efficient and safe tissue-selective targeted gene therapy approaches for delivering genes into keratocytes of the cornea in vivo using a normal or diseased rabbit model. New Zealand White rabbits, adeno-associated virus serotype 5 (AAV5), and a minimally invasive hair-dryer based vector-delivery technique were used. Fifty microliters of AAV5 titer (6.561012 vg/ml) expressing green fluorescent protein gene (GFP) was topically applied onto normal or diseased (fibrotic or neovascularized) rabbit corneas for 2-minutes with a custom vector-delivery technique. Corneal fibrosis and neovascularization in rabbit …

Microct Study Of Critical-Sized Defects Restored With A New Rhbmp-2 Carrier, Joanne S. Kim

Loma Linda University Electronic Theses, Dissertations & Projects

Introduction: Regenerating bone in critical-sized maxillofacial osseous defects remains a challenge. The utilization of recombinant human bone morphogenetic protein-2 (rhBMP-2) to form new bone has been shown to be a promising alternative to autogenous bone grafts, and the development of a carrier matrix to effectively deliver the rhBMP-2 has been identified as the critical factor necessary for the successful clinical application of rhBMP-2. The objective of this study was to perform a microCT analysis to examine the density of reconstructed bone using different rhBMP-2 carriers in critical-sized defects.

Material and Methods: Critical-sized defects (approximately 2.5 cm) were created in the …

Ricin B Chain-Insulin Fusion Protein Immunomodulation Of Type 1 Diabetes, James Edward Carter Iii

Loma Linda University Electronic Theses, Dissertations & Projects

Type 1 diabetes mellitus (T1D) is a debilitating chronic inflammatory disease of the insulin-producing pancreatic islet β-cells that results from a combination of genetic and environmental factors. Attempts to suppress Th1-mediated autoimmune diseases such as T1D by mucosal delivery of autoantigens for immunotolerization have yielded only partial success. Attainment of satisfactory levels of sustained immunological tolerance remains to be accomplished. To restore self-tolerance requires delivery of sufficient amounts of autoantigen to stimulate regulatory T helper cells that function to survey the gut and induce tolerance to consumed antigens such as food. Oral delivery of autoantigens has previously been shown to …

Gene Delivery In The Equine Cornea: A Novel Therapeutic Strategy, Dylan G. Buss, Ajay Sharma, Elizabeth A. Giuliano, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

Objective—To determine if hybrid adeno-associated virus serotype 2/5 (AAV5) vector can effectively deliver foreign genes into the equine cornea without causing adverse side effects. The aims of this study were to: (i) evaluate efficacy of AAV5 to deliver therapeutic genes into equine corneal fibroblasts (ECFs) using enhanced green fluorescent protein (EGFP) marker gene and (ii) establish the safety of AAV5 vector for equine corneal gene therapy.

Animal Material—Primary ECF cultures were harvested from healthy donor equine corneas. Cultures were maintained at 370C in humidified atmosphere with 5% CO2.

Procedure—AAV5 vector expressing EGFP under control of hybrid cytomegalovirus (CMV) + chicken …

Efficacy And Safety Of Mitomycin C As An Agent To Treat Corneal Scarring In Horses Using An In Vitro Model, Dylan G. Buss, Ajay Sharma, Elizabeth A. Giuliano, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

Objective—Mitomycin C (MMC) is used clinically to treat corneal scarring in human patients. We investigated the safety and efficacy of MMC to treat corneal scarring in horses by examining its effects at the early and late stages of disease using an in-vitro model.

Procedure—An in-vitro model of equine corneal fibroblast (ECF) developed was used. The equine corneal fibroblast or myofibroblast cultures were produced by growing primary ECF in the presence or absence of transforming growth factor beta-1 (TGFβ1) under serum-free conditions. The MMC dose for the equine cornea was defined with dose-dependent trypan blue exclusion and MTT [(3-4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assays …

Isolation And Cultivation Of Equine Corneal Keratocytes, Fibroblasts And Myofibroblasts, Dylan G. Buss, Elizabeth A. Giuliano, Ajay Sharma, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

Objective—To establish an in vitro model for the investigation of equine corneal wound healing. To accomplish this goal, a protocol to isolate and culture equine corneal keratocytes, fibroblasts and myofibroblasts was developed.

Animal material—Equine corneal buttons were aseptically harvested from healthy research horses undergoing humane euthanasia for reasons unrelated to this study. Slit-lamp biomicroscopy was performed prior to euthanasia by a board-certified veterinary ophthalmologist to ensure that all samples were harvested from horses free of anterior segment disease.

Procedure—Equine corneal stroma was isolated using mechanical techniques and stromal subsections were then cultured. Customized media at different culture conditions was used …

Vector Delivery Technique Affects Gene Transfer In The Cornea In Vivo, Rajiv R. Mohan, Ajay Sharma, Tyler C. Cebulko, Ashish Tandon

Pharmacy Faculty Articles and Research

Purpose: This study tested whether controlled drying of the cornea increases vector absorption in mouse and rabbit corneas in vivo and human cornea ex vivo, and studied the effects of corneal drying on gene transfer, structure and inflammatory reaction in the mouse cornea in vivo.

Methods: Female C57 black mice and New Zealand White rabbits were used for in vivo studies. Donor human corneas were used for ex vivo experiments. A hair dryer was used for drying the corneas after removing corneal epithelium by gentle scraping. The corneas received no, once, twice, thrice, or five times warm air for …

Localization Of Angiotensin Converting Enzyme In Rabbit Cornea And Its Role In Controlling Corneal Angiogenesis In Vivo, Ajay Sharma, Daniel I. Bettis, John W. Cowden, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

Purpose: The renin angiotensin system (RAS) has been shown to modulate vascular endothelial growth factor and angiogenesis. In this study we investigated (i) the existence of the RAS components angiotensin converting enzyme (ACE) and angiotensin II receptors (AT1 and AT2) in the rabbit cornea using in vitro and ex vivo models and (ii) the effect of enalapril, an ACE inhibitor, to inhibit angiogenesis in rabbit cornea in vivo.

Methods: New Zealand White rabbits were used. Cultured corneal fibroblasts and corneal epithelial cells were used for RNA isolation and cDNA preparation using standard molecular biology techniques. PCR was performed to …

Aav Serotype Influences Gene Transfer In Corneal Stroma In Vivo, Ajay Sharma, Jonathan C. K. Tovey, Arkasubhra Ghosh, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

This study evaluated the cellular tropism and relative transduction efficiency of three AAV serotypes, AAV6, AAV8 and AAV9, for corneal gene delivery using mouse cornea in vivo and donor human cornea ex vivo. The AAV6, AAV8 and AAV9 serotypes having AAV2 plasmid encoding for alkaline phosphatase (AP) gene were generated by transfecting HEK293 cell line with pHelper, pARAP4 and pRep/Cap plasmids. Viral vectors (109 vg/μl) were topically applied onto mouse cornea in vivo and human cornea ex vivo after removing the epithelium. Human corneas were processed for transgene delivery at day 5 after viral vector application. Mouse corneas were harvested …

Mnte-2-Pyp And Radiation In A Prostate Cancer Model: Implications For Radiotherapy, Adeola Y. Makinde

Loma Linda University Electronic Theses, Dissertations & Projects

A major limitation of successful radiation therapy in cancer treatment is the increase in normal tissue damage as higher doses are used to achieve greater tumor destruction. Radiation dose optimization in cancer therapy requires achieving maximum tumor destruction with minimal damage to normal tissue Antioxidants have been shown to protect normal tissues against radiation damage, as radiation-induced tissue damage results predominantly from reactive oxygen species that directly damage cellular components. However, for effective use as normal tissue radioprotectants in radiotherapy, these antioxidants must not protect the tumors. Mn (III) tetrakis (N-ethylpyridinium-2-yl) porphyrin (MnTE-2-PyP) is a metalloporphyrin antioxidant that has been …

Trichostatin A Inhibits Corneal Haze In Vitro And In Vivo, Ajay Sharma, Maneesh M. Mehan, Sunilima Sinha, John W. Cowden, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

PURPOSE. Trichostatin A (TSA), a histone deacetylase inhibitor, has been shown to suppress TGF- –induced fibrogenesis in many nonocular tissues. The authors evaluated TSA cytotoxicity and its antifibrogenic activity on TGF- –driven fibrosis in the cornea with the use of in vitro and in vivo models.

METHODS. Human corneal fibroblasts (HSFs) were used for in vitro studies, and New Zealand White rabbits were used for in vivo studies. Haze in the rabbit cornea was produced with photorefractive keratectomy (PRK) using excimer laser. Trypan blue exclusion and MTT assays evaluated TSA cytotoxicity to the cornea. Density of haze in the rabbit …

Anti-Inflammatory Effects Of Ppar-Gamma In Surgical Brain Injury (Sbi), Amy Hyong

Loma Linda University Electronic Theses, Dissertations & Projects

Introduction: Brain injury can be caused by neurosurgical procedures themselves, due to direct trauma, retractor stretch, intraoperative hemorrhage and electrocautery damage. As a result of this surgical brain injury (SBI) postoperative complications such as inflammation, brain edema, and cell death can occur in the susceptible brain areas. Cerebral inflammation is a known contributor to the pathophysiology of brain injury. Following brain injury, the release of inflammatory mediators facilitates the development of BBB breakdown, cerebral edema, oxidative stress and neuronal death, resulting in further tissue damage in the brain and poor neurological outcomes. This study evaluates whether the use of a …