Delivery of PAMAM dendrimers and dendriplexes across natural barriers (blood-brain barrier and placental barrier) in healthy pregnant mice.

Gene therapy is an important tool for treating fetal diseases that allows for the delivery and integration of therapeutic genes into the genome of cells carrying mutations. Nanomolecules, like PAMAM dendrimers, have recently come into wider use for carrying vectors as they have several advantages over viral vectors. Namely, (1) tunable size and surface chemistry, (2) uniform size, (3) the ability to target specific tissues, and (4) the ability to carry large biomolecules and drugs. Recently, we demonstrated that 4th generation (G4) PAMAM dendrimer with a cystamine core and a non-toxic surface having 90% -OH and 10% -NH2 groups (D-Cys) could cross the blood-brain barrier following injection into the bloodstream. In the current study, as a proof of concept, we delivered the dendrimers alone (D-Cys) tagged with Cy5.5 (D-Cys-cy5.5) to healthy pregnant C57BL/6J mice to determine the fate of these dendrimers in the pregnant mice as well as in the fetus. Systematic diffusion of the D-Cys-cy5.5 was evaluated on gestational day 17 (3 days after injection) using in vivo imaging. This revealed that the dendrimer was taken up into circulation and away from the injection site. Analysis of sections by fluorescence microscopy showed that D-Cys-cy5.5 was able to successfully cross the maternal blood-brain barrier. However, analysis of the fetal brains failed to detect dendrimers in the central nervous system (CNS). Instead, they appeared to be retained in the placenta. This is one of the first studies to analyze the distribution of surface-modified PAMAM dendrimer in the pregnant mouse and fetus following systemic injection.

Unilateral Administration of Surface-Modified G1 and G4 PAMAM Dendrimers in Healthy Mice to Assess Dendrimer Migration in the Brain.

Polyamidoamine (PAMAM) dendrimers are nanoparticles that have a wide scope in the field of biomedicine. Previous evidence shows that the generation 4 (G4) dendrimers with a 100% amine surface (G4-NH2) are highly toxic to cells in vitro and in vivo due to their positively charged amine groups. To reduce the toxicity, we modified the surface of the dendrimers to have more neutral functional groups, with 10% of the surface covered with -NH2 and 90% of the surface covered with hydroxyl groups (-OH; G4-90/10). Our previous in vitro data show that these modified dendrimers are taken up by cells, neurons, and different types of stem cells in vitro and neurons and glial cells in vivo. The toxicity assay shows that these modified dendrimers are less toxic compared with G4-NH2 dendrimers. Moreover, prolonged dendrimer exposure (G1-90/10 and G4-90/10), up to 3 weeks following unilateral intrastriatal injections into the striatum of mice, showed that dendrimers have the tendency to migrate within the brain via corpus callosum at different rates depending on their size. We also found that there is a difference in migration between the G1 and G4 dendrimers based on their size differences. The G4 dendrimers migrate in the anterior and posterior directions as well as more laterally from the site of injection in the striatum compared to the G1 dendrimers. Moreover, the G4 dendrimers have unique projections from the site of injection to the cortical areas.

Nanoparticle-Based Therapies in Hypertension.

Nearly 1.4 billion people worldwide suffer from arterial hypertension, a significant risk factor for cardiovascular disease which is now the leading cause of death. Despite numerous drugs designed to treat hypertension, only ≈14% of hypertensive individuals have their blood pressure under control. A critical factor negatively impacting the efficacy of available treatments is their poor bioavailability. This leads to increased dosing requirements which can result in more side effects, resulting in patient noncompliance. A recent solution to improve dosing and bioavailability issues has been to incorporate drugs into nanoparticle carriers, with over 50 nanodrugs currently on the market across all diseases, and another 51 currently in clinical trials. Given their ability to improve solubility and bioavailability, nanoparticles may offer significant advantages in the formulation of antihypertensives to overcome pharmacokinetic shortcomings. To date, however, no antihypertensive nanoformulations have been clinically approved. This review assesses in vivo study data from preclinical antihypertensive nanoformulation development and testing. Combined, the results of these studies suggest nanoformulation of antihypertensive drugs may be a promising solution to overcome the poor efficacy of currently available antihypertensives, and with further advances has the potential to open paths for new substances that have heretofore been clinically unrealistic due to poor bioavailability.

Dual pH and microbial-sensitive galactosylated polymeric nanocargoes for multi-level targeting to combat ulcerative colitis.

Ulcerative colitis (UC) is a type of inflammatory bowel disease characterized by inflammation, ulcers and irritation of the mucosal lining. Oral drug delivery in UC encounters challenges because of multifaceted barriers. Dexamethasone-loaded galactosylated-PLGA/Eudragit S100/pullulan nanocargoes (Dexa-GP/ES/Pu NCs) have been developed with a dual stimuli-sensitive coating responsive to both colonic pH and microbiota, and an underneath galactosylated-PLGA core (GP). The galactose ligand of the GP preferentially binds to the macrophage galactose type-lectin-C (MGL-2) surface receptor. Therefore, both stimuli and ligand-mediated targeting facilitate nanocargoes to deliver Dexa specifically to the colon with enhanced macrophage uptake. Modified emulsion method coupled with a solvent evaporation coating technique was employed to prepare Dexa-GP/ES/Pu NCs. The nanocargoes were tested using in vitro, ex vivo techniques and dextran sodium sulfate (DSS) induced UC model. Prepared nanocargoes had desired physicochemical properties, drug release, cell uptake and cellular viability. Investigations using a DSS-colitis model showed high localization and mitigation of colitis with downregulation of NF-ĸB and COX-2, and restoration of clinical, histopathological, biochemical indices, antioxidant balance, microbial alterations, FTIR spectra, and epithelial junctions' integrity. Thus, Dexa-GP/ES/Pu NCs found to be biocompatible nanocargoes capable of delivering drugs to the inflamed colon with unique targeting properties for prolonged duration.

Housing R6/2 Mice with Wild-Type Littermates Increases Lifespan.

The R6/2 murine model of Huntington's disease (HD) is extensively used in HD research. The current study replicates and extends previous work assessing the impact of housing R6/2 mice with healthy wild-type (WT) littermates on disease progression. The current study extends the previous finding by including male cohorts and the use of a standard diet and water regimen, as opposed to the enhanced diet used in the previous study. This study found that the inclusion of healthy wild-type (WT) littermates, alone, improved survivabilty in R6/2 mice, but did not have a significant impact on weight loss.

Solid Lipid Curcumin Particles Protect Medium Spiny Neuronal Morphology, and Reduce Learning and Memory Deficits in the YAC128 Mouse Model of Huntington's Disease.

Huntington's disease (HD) is a genetic neurodegenerative disorder characterized by motor, cognitive, and psychiatric symptoms, accompanied by massive neuronal degeneration in the striatum. In this study, we utilized solid lipid curcumin particles (SLCPs) and solid lipid particles (SLPs) to test their efficacy in reducing deficits in YAC128 HD mice. Eleven-month-old YAC128 male and female mice were treated orally with SLCPs (100 mg/kg) or equivalent volumes of SLPs or vehicle (phosphate-buffered saline) every other day for eight weeks. Learning and memory performance was assessed using an active-avoidance task on week eight. The mice were euthanized, and their brains were processed using Golgi-Cox staining to study the morphology of medium spiny neurons (MSNs) and Western blots to quantify amounts of DARPP-32, brain-derived neurotrophic factor (BDNF), TrkB, synaptophysin, and PSD-95. We found that both SLCPs and SLPs improved learning and memory in HD mice, as measured by the active avoidance task. We also found that SLCP and SLP treatments preserved MSNs arborization and spinal density and modulated synaptic proteins. Our study shows that SLCPs, as well as the lipid particles, can have therapeutic effects in old YAC128 HD mice in terms of recovering from HD brain pathology and cognitive deficits.

Latency to startle is increased in the 5xFAD mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that results in cognitive decline and a number of other neuropsychiatric symptoms. One area that is often affected by neuropsychiatric disease is the response to sudden, loud noises, as measured by the acoustic startle response (ASR), and prepulse inhibition (PPI), which indicates sensory-gating abilities. Evidence suggests AD patients, even early in the disease, show alteration in ASR. Studies have also shown changes in this measure in transgenic mouse models of AD. To assess the homology of 5xFAD mice to AD patients, the current study analyzed several aspects of the startle response in these mice using a protocol with fewer trials than previous studies. It was found that the 5xFAD mice had a delayed startle response, similar to what has been observed in AD sufferers. These results suggest the ASR may be a useful tool in assessing the efficacy of potential therapeutics, and that a simplified protocol may be more sensitive to between-groups differences for this task.

Diabetes and Associated Cardiovascular Complications in American Indians/Alaskan Natives: A Review of Risks and Prevention Strategies.

Diabetes mellitus (DM) is the seventh leading cause of death in the United States and the leading cause of death in the U.S. American Indian/Alaskan Natives (AI/ANs), who comprise only 2% of the total population. The AI/AN population has a high prevalence of DM in adults aged 20 years or older and is developing DM at a younger age than the general U.S. POPULATION: DM is a major risk factor for cardiovascular disease (CVD), and mortality from CVD is higher in AI/ANs than the general population, as is the prevalence of stroke and 1-year poststroke mortality for both genders when compared to non-Hispanic whites. A genome-wide scan found a number of chromosome linkages in the AI/AN population that suggest that genetic factors may contribute to their high risk of DM and CVD. Importantly, studies also suggest that in addition to race/ethnicity, cultural norms and historic conditions play important roles in the prevalence of DM and CVD in this population. Therefore, multiple factors should be taken into consideration when establishing prevention programs to decrease the prevalence of obesity, diabetes, and CVD incidence among adults and children in the AI/AN population. Prevention programs should focus on behavioral risk factors and lifestyle changes like encouraging smoking cessation, healthy diet, and increased physical activity while taking into consideration cultural, economic, and geographic factors.

Metformin: An Old Drug with New Applications.

Metformin is a biguanide drug that has been used to treat type 2 diabetes mellitus for more than 60 years. The United Kingdom Prospective Diabetic Study (UKPDS) has shown metformin to improve mortality rates in diabetes patients, and recent studies suggest metformin has additional effects in treating cancer, obesity, nonalcoholic fatty liver disease (NAFLD), polycystic ovary syndrome (PCOS), and metabolic syndrome. Metformin has also been shown to alleviate weight gain associated with antipsychotic medication. Metformin has recently been extensively studied and emerging evidence suggests metformin decreases hepatocyte triglyceride accumulation in NAFLD and prevents liver tumorigenesis. Interestingly, studies have also shown metformin reduces visceral fat, suppresses white-adipose-tissue (WAT) extracellular matrix remodeling, and inhibits obesity-induced inflammation. However, clinical evidence for using metformin to treat NAFLD, cancer, metabolic syndrome, or to prevent hepatocellular carcinoma in NAFLD patients is lacking. This review therefore addresses the potential beneficial effects of metformin on NAFLD, its role in protecting against cardiac ischemia⁻reperfusion (I/R) injury, atherosclerosis, glucotoxicity, and lipotoxicity induced oxidative and ER stress in pancreatic ß-cell dysfunction, as well as its underlying molecular mechanisms of action.

Induced Pluripotent Stem Cell-Derived Neural Stem Cell Transplantations Reduced Behavioral Deficits and Ameliorated Neuropathological Changes in YAC128 Mouse Model of Huntington's Disease.

Huntington's disease (HD) is a genetic neurodegenerative disorder characterized by neuronal loss and motor dysfunction. Although there is no effective treatment, stem cell transplantation offers a promising therapeutic strategy, but the safety and efficacy of this approach needs to be optimized. The purpose of this study was to test the potential of intra-striatal transplantation of induced pluripotent stem cell-derived neural stem cells (iPS-NSCs) for treating HD. For this purpose, we developed mouse adenovirus-generated iPSCs, differentiated them into neural stem cells in vitro, labeled them with Hoechst, and transplanted them bilaterally into striata of 10-month old wild type (WT) and HD YAC128 mice. We assessed the efficiency of these transplanted iPS-NSCs to reduce motor deficits in YAC128 mice by testing them on an accelerating rotarod task at 1 day prior to transplantation, and then weekly for 10 weeks. Our results showed an amelioration of locomotor deficits in YAC128 mice that received iPS-NSC transplantations. Following testing, the mice were sacrificed, and their brains were analyzed using immunohistochemistry and Western blot (WB). The results from our histological examinations revealed no signs of tumors and evidence that many iPS-NSCs survived and differentiated into region-specific neurons (medium spiny neurons) in both WT and HD mice, as confirmed by co-labeling of Hoechst-labeled transplanted cells with NeuN and DARPP-32. Also, counts of Hoechst-labeled cells revealed that a higher proportion were co-labeled with DARPP-32 and NeuN in HD-, compared to WT- mice, suggesting a dissimilar differentiation pattern in HD mice. Whereas significant decreases were found in counts of NeuN- and DARPP-32-labeled cells, and for neuronal density measures in striata of HD vehicle controls, such decrements were not observed in the iPS-NSCs-transplanted-HD mice. WB analysis showed increase of BDNF and TrkB levels in striata of transplanted HD mice compared to HD vehicle controls. Collectively, our data suggest that iPS-NSCs may provide an effective option for neuronal replacement therapy in HD.

Intrastriatal transplantation of adenovirus-generated induced pluripotent stem cells for treating neuropathological and functional deficits in a rodent model of Huntington's disease.

Induced pluripotent stem cells (iPSCs) show considerable promise for cell replacement therapies for Huntington's disease (HD). Our laboratory has demonstrated that tail-tip fibroblasts, reprogrammed into iPSCs via two adenoviruses, can survive and differentiate into neuronal lineages following transplantation into healthy adult rats. However, the ability of these cells to survive, differentiate, and restore function in a damaged brain is unknown. To this end, adult rats received a regimen of 3-nitropropionic acid (3-NP) to induce behavioral and neuropathological deficits that resemble HD. At 7, 21, and 42 days after the initiation of 3-NP or vehicle, the rats received intrastriatal bilateral transplantation of iPSCs. All rats that received 3-NP and vehicle treatment displayed significant motor impairment, whereas those that received iPSC transplantation after 3-NP treatment had preserved motor function. Histological analysis of the brains of these rats revealed significant decreases in optical densitometric measures in the striatum, lateral ventricle enlargement, as well as an increase in striosome size in all rats receiving 3-NP when compared with sham rats. The 3-NP-treated rats given transplants of iPSCs in the 7- or 21-day groups did not exhibit these deficits. Transplantation of iPSCs at the late-stage (42-day) time point did not protect against the 3-NP-induced neuropathology, despite preserving motor function. Transplanted iPSCs were found to survive and differentiate into region-specific neurons in the striatum of 3-NP rats, at all transplantation time points. Taken together, these results suggest that transplantation of adenovirus-generated iPSCs may provide a potential avenue for therapeutic treatment of HD.

Spatial learning deficits and emotional impairments in pentylenetetrazole-kindled rats.

Pentylenetetrazole (PTZ) is a chemical kindling agent used to examine the efficacy of potential anticonvulsants in rats. However, the extent to which PTZ mimics postseizure symptoms of epilepsy has not been thoroughly examined. This study assessed whether PTZ-induced seizures produce cognitive and emotional deficits that mimic those observed in many epileptic patients. Rats were given 30mg/kg PTZ or vehicle (intraperitoneally) every other day for 28 days. Those rats exhibiting consistent seizure activity were tested for learning ability and emotional reactivity, beginning 1 week following a single challenge dose of PTZ. Rats given PTZ made more reference memory errors in a radial arm water maze task, and exhibited emotional abnormalities in the forced swim test, the systematic handling test, and the open-field exploratory maze. Histological analysis revealed neuronal loss in the CA1 area and increased mossy fiber sprouting in the dentate gyrus, similar to what is observed in human epilepsy. These results indicate that PTZ kindling provides a useful model of postseizure dysfunction, which can serve as a screen for potential treatments for those cognitive, emotional, and neuropathological deficits that resemble those symptoms observed in human epilepsy.