Multifaceted Mechanisms of Vascular Calcification in Aging.

Approximately 20% of the world's population will be around or above 65 years of age by the next decade. Out of these, 40% are suspected to have cardiovascular diseases as a cause of mortality. Arteriosclerosis, characterized by increased vascular calcification, impairing Windkessel effect and tissue perfusion, and determining end-organ damage, is a hallmark of vascular pathology in the elderly population. Risk factors accumulated during aging affect the normal physiological and vascular aging process, which contributes to the progression of arteriosclerosis. Traditional risk factors, age-associated diseases, and respective regulating mechanisms influencing vascular calcification and vascular stiffness have been extensively studied for many years. Despite the well-known fact that aging alone can induce vascular damage, specific mechanisms that implicate physiological aging in vascular calcification, contributing to vascular stiffness, are poorly understood. This review focuses on mechanisms activated during normal aging, for example, cellular senescence, autophagy, extracellular vesicles secretion, and oxidative stress, along with the convergence of premature aging models' pathophysiology, such as Hutchinson-Gilford Progeria (prelamin accumulation) and Klotho deficiency, to understand vascular calcification in aging. Understanding the mechanisms of vascular damage in aging that intersect with age-associated diseases and risk factors is crucial to foster innovative therapeutic targets to mitigate cardiovascular disease. Visual Overview- An online visual overview is available for this article.

Expansive Vascular Remodeling and Increased Vascular Calcification Response to Cholecalciferol in a Murine Model of Obesity and Insulin Resistance.

Objective- We hypothesized that ob/ob mice develop expansive vascular remodeling associated with calcification. Approach and Results- We quantified and investigated mechanisms of vascular remodeling and vascular calcification in ob/ob mice after vitamin D3(VD) stimulation or PBS (control), compared with C57BL/6 mice. Both ob/ob (OBVD [VD-treated ob/ob mice]) and C57BL/6 (C57VD [VD-treated C57BL/6 mice]) received 8×103 IU/day of intraperitoneal VD for 14 days. Control ob/ob (OBCT [PBS-treated ob/ob mice]) and C57BL/6 (C57CT [PBS-treated C57BL/6 mice]) received intraperitoneal PBS for 14 days. Hypervitaminosis D increased the external and internal elastic length in aortae from OBVD, resulting in increased total vascular area and lumen vascular area, respectively, which characterizes expansive vascular remodeling. OBVD decreased the aortic wall thickness, resulting in hypotrophic vascular remodeling. We demonstrated increased collagen deposition, elastolysis, and calcification in aortae from OBVD. Our results showed a positive correlation between expansive vascular remodeling and vascular calcification in OBVD. We demonstrated increased serum calcium levels, augmented Bmp (bone morphogenetic protein)-2 and osteochondrogenic proteins expression in OBVD aortae. Furthermore, aortae from OBVD increased oxidative stress, coincidently with augmented in situ MMP (matrix metalloproteinase) activity and exhibited no VDR (VD receptor) inhibition after VD. Conclusions- Our data provide evidence that obese and insulin-resistant mice (ob/ob) developed expansive hypotrophic vascular remodeling correlated directly with increased vascular calcification after chronic VD stimulation. Positive hypotrophic vascular remodeling and vascular calcification in this mouse model is possibly mediated by the convergence of absence VDR downregulation after VD stimulation, increased reactive oxygen species generation, and MMP activation.

Therapeutic Efficiency of Multiple Applications of Magnetic Hyperthermia Technique in Glioblastoma Using Aminosilane Coated Iron Oxide Nanoparticles: In Vitro and In Vivo Study.

Magnetic hyperthermia (MHT) has been shown as a promising alternative therapy for glioblastoma (GBM) treatment. This study consists of three parts: The first part evaluates the heating potential of aminosilane-coated superparamagnetic iron oxide nanoparticles (SPIONa). The second and third parts comprise the evaluation of MHT multiple applications in GBM model, either in vitro or in vivo. The obtained heating curves of SPIONa (100 nm, +20 mV) and their specific absorption rates (SAR) stablished the best therapeutic conditions for frequencies (309 kHz and 557 kHz) and magnetic field (300 Gauss), which were stablished based on three in vitro MHT application in C6 GBM cell line. The bioluminescence (BLI) signal decayed in all applications and parameters tested and 309 kHz with 300 Gauss have shown to provide the best therapeutic effect. These parameters were also established for three MHT applications in vivo, in which the decay of BLI signal correlates with reduced tumor and also with decreased tumor glucose uptake assessed by positron emission tomography (PET) images. The behavior assessment showed a slight improvement after each MHT therapy, but after three applications the motor function displayed a relevant and progressive improvement until the latest evaluation. Thus, MHT multiple applications allowed an almost total regression of the GBM tumor in vivo. However, futher evaluations after the therapy acute phase are necessary to follow the evolution or tumor total regression. BLI, positron emission tomography (PET), and spontaneous locomotion evaluation techniques were effective in longitudinally monitoring the therapeutic effects of the MHT technique.

Methods of Granulocyte Isolation from Human Blood and Labeling with Multimodal Superparamagnetic Iron Oxide Nanoparticles.

This in vitro study aimed to find the best method of granulocyte isolation for subsequentlabeling with multimodal nanoparticles (magnetic and fluorescent properties) to enable detectionby optical and magnetic resonance imaging (MRI) techniques. The granulocytes were obtained fromvenous blood samples from 12 healthy volunteers. To achieve high purity and yield, four differentmethods of granulocyte isolation were evaluated. The isolated granulocytes were labeled withmultimodal superparamagnetic iron oxide nanoparticles (M-SPIONs) coated with dextran, and theiron load was evaluated qualitatively and quantitatively by MRI, near-infrared fluorescence (NIRF)and inductively coupled plasma mass spectrometry (ICP-MS). The best method of granulocyteisolation was Percoll with Ficoll, which showed 95.92% purity and 94% viability. After labeling withM-SPIONs, the granulocytes showed 98.0% purity with a yield of 3.5 × 106 cells/mL and more than98.6% viability. The iron-loading value in the labeled granulocytes, as obtained by MRI, was 6.40 ±0.18 pg/cell. Similar values were found with the ICP-MS and NIRF imaging techniques. Therefore,our study shows that it is possible to isolate granulocytes with high purity and yield and labelingwith M-SPIONs provides a high internalized iron load and low toxicity to cells. Therefore, these MSPION-labeled granulocytes could be a promising candidate for future use ininflammation/infection detection by optical and MRI techniques.

The Effects of Omega-3 Polyunsaturated Fatty Acids on Breast Cancer as a Preventive Measure or as an Adjunct to Conventional Treatments.

In order to understand how omega-3 polyunsaturated fatty acid (ω-3 PUFA) supplements affect breast cancer prevention and treatment, a systematic review of articles published in the last 5 years in two databases was performed. Of the 679 articles identified, only 27 were included and examined based on five topics, taking into account: the induction type of the breast cancer used in animal models; the characteristics of the induction model by cell transplantation; the experimental design of the ω-3 supplementation-combined or not with a treatment antitumor drug; the fatty acids (FAs) composition used; the analysis of the studies' outcomes. There are diverse and well-established animal models of breast cancer in the literature, with very relevant histological and molecular similarities depending on the specific objective of the study, such as whether the method of tumor induction was transgenic, by cell transplantation, or by oncogenic drugs. The analyses of outcomes were mainly focused on monitoring tumor growth, body/tumor weight, and molecular, genetic, or histological analyses, and few studies evaluated latency, survival, or metastases. The best results occurred when supplementation with ω-3 PUFA was associated with antitumor drugs, especially in the analysis of metastases and volume/weight of tumors or when the supplementation was started early and maintained for a long time. However, the beneficial effect of ω-3 PUFA supplementation when not associated with an antitumor agent remains unclear.

Bioluminescence Imaging and ICP-MS Associated with SPION as a Tool for Hematopoietic Stem and Progenitor Cells Homing and Engraftment Evaluation.

Bone marrow transplantation is a treatment for a variety of hematological and non-hematological diseases. For the transplant success, it is mandatory to have a thriving engraftment of transplanted cells, which directly depends on their homing. The present study proposes an alternative method to evaluate the homing and engraftment of hematopoietic stem cells using bioluminescence imaging and inductively coupled plasma mass spectrometry (ICP-MS) associated with superparamagnetic iron oxide nanoparticles. We have identified an enriched population of hematopoietic stem cells in the bone marrow following the administration of Fluorouracil (5-FU). Lately, the cell labeling with nanoparticles displayed the greatest internalization status when treated with 30 µg Fe/mL. The quantification by ICP-MS evaluate the stem cells homing by identifying 3.95 ± 0.37 µg Fe/mL in the control and 6.61 ± 0.84 µg Fe/mL in the bone marrow of transplanted animals. In addition, 2.14 ± 0.66 mg Fe/g in the spleen of the control group and 2.17 ± 0.59 mg Fe/g in the spleen of the experimental group was also measured. Moreover, the bioluminescence imaging provided the follow up on the hematopoietic stem cells behavior by monitoring their distribution by the bioluminescence signal. Lastly, the blood count enabled the monitoring of animal hematopoietic reconstitution and ensured the transplantation effectiveness.

Superparamagnetic and highly bioactive SPIONS/bioactive glass nanocomposite and its potential application in magnetic hyperthermia.

Magnetic bioactive glass-ceramics are biomaterials applied for magnetic hyperthermia in bone cancer treatment, thereby treating the bone tumor besides regenerating the damaged bone. However, combining high bioactivity and high saturation magnetization remains a challenge since the thermal treatment step employed to grow magnetic phases is also related to loss of bioactivity. Here, we propose a new nanocomposite made of superparamagnetic iron oxide nanoparticles (SPIONs) dispersed in a sol-gel-derived bioactive glass matrix, which does not need any thermal treatment for crystallization of magnetic phases. The scanning and transmission electron microscopies, X-ray diffraction, and dynamic light scattering results confirm that the SPIONs are actually embedded in a nanosized glass matrix, thus forming a nanocomposite. Magnetic and calorimetric characterizations evidence their proper behavior for hyperthermia applications, besides evidencing inter-magnetic nanoparticle interactions within the nanocomposite. Bioactivity and in vitro characterizations show that such nanocomposites exhibit apatite-forming properties similar to the highly bioactive parent glass, besides being osteoinductive. This methodology is a new alternative to produce magnetic bioactive materials to which the magnetic properties only rely on the quality of the SPIONs used in the synthesis. Thereby, these nanocomposites can be recognized as a new class of bioactive materials for applications in bone cancer treatment by hyperthermia.

Optimization of Multimodal Nanoparticles Internalization Process in Mesenchymal Stem Cells for Cell Therapy Studies.

Considering there are several difficulties and limitations in labeling stem cells using multifunctional nanoparticles (MFNP), the purpose of this study was to determine the optimal conditions for labeling human bone marrow mesenchymal stem cells (hBM-MSC), aiming to monitor these cells in vivo. Thus, this study provides information on hBM-MSC direct labeling using multimodal nanoparticles in terms of concentration, magnetic field, and period of incubation while maintaining these cells' viability and the homing ability for in vivo experiments. The cell labeling process was assessed using 10, 30, and 50 µg Fe/mL of MFNP, with periods of incubation ranging from 4 to 24 h, with or without a magnetic field, using optical microscopy, near-infrared fluorescence (NIRF), and inductively coupled plasma mass spectrometry (ICP-MS). After the determination of optimal labeling conditions, these cells were applied in vivo 24 h after stroke induction, intending to evaluate cell homing and improve NIRF signal detection. In the presence of a magnetic field and utilizing the maximal concentration of MFNP during cell labeling, the iron load assessed by NIRF and ICP-MS was four times higher than what was achieved before. In addition, considering cell viability higher than 98%, the recommended incubation time was 9 h, which corresponded to a 25.4 pg Fe/cell iron load (86% of the iron load internalized in 24 h). The optimization of cellular labeling for application in the in vivo study promoted an increase in the NIRF signal by 215% at 1 h and 201% at 7 h due to the use of a magnetized field during the cellular labeling process. In the case of BLI, the signal does not depend on cell labeling showing no significant differences between unlabeled or labeled cells (with or without a magnetic field). Therefore, the in vitro cellular optimized labeling process using magnetic fields resulted in a shorter period of incubation with efficient iron load internalization using higher MFNP concentration (50 µgFe/mL), leading to significant improvement in cell detection by NIRF technique without compromising cellular viability in the stroke model.

The Advances in Glioblastoma On-a-Chip for Therapy Approaches.

This systematic review aimed to verify the use of microfluidic devices in the process of implementing and evaluating the effectiveness of therapeutic approaches in glioblastoma on-a-chip, providing a broad view of advances to date in the use of this technology and their perspectives. We searched studies with the variations of the keywords "Glioblastoma", "microfluidic devices", "organ-on-a-chip" and "therapy" of the last ten years in PubMed and Scopus databases. Of 446 articles identified, only 22 articles were selected for analysis according to the inclusion and exclusion criteria. The microfluidic devices were mainly produced by soft lithography technology, using the PDMS material (72%). In the microenvironment, the main extracellular matrix used was collagen type I. Most studies used U87-MG glioblastoma cells from humans and 31.8% were co-cultivated with HUVEC, hCMEC/D3, and astrocytes. Chemotherapy was the majority of therapeutic approaches, assessing mainly the cellular viability and proliferation. Furthermore, some alternative therapies were reported in a few studies (22.6%). This study identified a diversity of glioblastoma on-a-chip to assess therapeutic approaches, often using intermediate levels of complexity. The most advanced level implemented the intersection between different biological systems (liver-brain or intestine-liver-brain), BBB model, allowing in vitro studies with greater human genetic similarity, reproducibility, and low cost, in a highly customizable platform.

Advances in Concentration Gradient Generation Approaches in a Microfluidic Device for Toxicity Analysis.

This systematic review aimed to analyze the development and functionality of microfluidic concentration gradient generators (CGGs) for toxicological evaluation of different biological organisms. We searched articles using the keywords: concentration gradient generator, toxicity, and microfluidic device. Only 33 of the 352 articles found were included and examined regarding the fabrication of the microdevices, the characteristics of the CGG, the biological model, and the desired results. The main fabrication method was soft lithography, using polydimethylsiloxane (PDMS) material (91%) and SU-8 as the mold (58.3%). New technologies were applied to minimize shear and bubble problems, reduce costs, and accelerate prototyping. The Christmas tree CGG design and its variations were the most reported in the studies, as well as the convective method of generation (61%). Biological models included bacteria and nematodes for antibiotic screening, microalgae for pollutant toxicity, tumor and normal cells for, primarily, chemotherapy screening, and Zebrafish embryos for drug and metal developmental toxicity. The toxic effects of each concentration generated were evaluated mostly with imaging and microscopy techniques. This study showed an advantage of CGGs over other techniques and their applicability for several biological models. Even with soft lithography, PDMS, and Christmas tree being more popular in their respective categories, current studies aim to apply new technologies and intricate architectures to improve testing effectiveness and reduce common microfluidics problems, allowing for high applicability of toxicity tests in different medical and environmental models.

Effect of Cell Therapy and Exercise Training in a Stroke Model, Considering the Cell Track by Molecular Image and Behavioral Analysis.

The goal of this study is to see how combining physical activity with cell treatment impacts functional recovery in a stroke model. Molecular imaging and multimodal nanoparticles assisted in cell tracking and longitudinal monitoring (MNP). The viability of mesenchymal stem cell (MSC) was determined using a 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay and bioluminescent image (BLI) after lentiviral transduction and MNP labeling. At random, the animals were divided into 5 groups (control-G1, and experimental G2-G5). The photothrombotic stroke induction was confirmed by local blood perfusion reduction and Triphenyltetrazolium chloride (TTC), and MSC in the G3 and G5 groups were implanted after 24 h, with BLI and near-infrared fluorescence image (NIRF) tracking these cells at 28 h, 2, 7, 14, and 28 days. During a 28-day period, the G5 also conducted physical training, whereas the G4 simply did the training. At 0, 7, 14, and 28 days, the animals were functionally tested using a cylinder test and a spontaneous motor activity test. MNP internalization in MSC was confirmed using brightfield and fluorescence microscopy. In relation to G1 group, only 3% of cell viability reduced. The G2-G5 groups showed more than 69% of blood perfusion reduction. The G5 group performed better over time, with a progressive recovery of symmetry and an increase of fast vertical movements. Up to 7 days, BLI and NIRF followed MSC at the damaged site, demonstrating a signal rise that could be connected to cell proliferation at the injury site during the acute phase of stroke. Local MSC therapy mixed with physical activity resulted in better results in alleviating motor dysfunction, particularly during the acute period. When it comes to neurorehabilitation, this alternative therapy could be a suitable fit.

LN-Derived Fibroblastic Reticular Cells and Their Impact on T Cell Response-A Systematic Review.

Fibroblastic reticular cells (FRCs), usually found and isolated from the T cell zone of lymph nodes, have recently been described as much more than simple structural cells. Originally, these cells were described to form a conduit system called the "reticular fiber network" and for being responsible for transferring the lymph fluid drained from tissues through afferent lymphatic vessels to the T cell zone. However, nowadays, these cells are described as being capable of secreting several cytokines and chemokines and possessing the ability to interfere with the immune response, improving it, and also controlling lymphocyte proliferation. Here, we performed a systematic review of the several methods employed to investigate the mechanisms used by fibroblastic reticular cells to control the immune response, as well as their ability in determining the fate of T cells. We searched articles indexed and published in the last five years, between 2016 and 2020, in PubMed, Scopus, and Cochrane, following the PRISMA guidelines. We found 175 articles published in the literature using our searching strategies, but only 24 articles fulfilled our inclusion criteria and are discussed here. Other articles important in the built knowledge of FRCs were included in the introduction and discussion. The studies selected for this review used different strategies in order to access the contribution of FRCs to different mechanisms involved in the immune response: 21% evaluated viral infection in this context, 13% used a model of autoimmunity, 8% used a model of GvHD or cancer, 4% used a model of Ischemic-reperfusion injury (IRI). Another four studies just targeted a particular signaling pathway, such as MHC II expression, FRC microvesicles, FRC secretion of IL-15, FRC network, or ablation of the lysophosphatidic acid (LPA)-producing ectoenzyme autotaxin. In conclusion, our review shows the strategies used by several studies to isolate and culture fibroblastic reticular cells, the models chosen by each one, and dissects their main findings and implications in homeostasis and disease.

In Vitro Evaluation of Hyperthermia Magnetic Technique Indicating the Best Strategy for Internalization of Magnetic Nanoparticles Applied in Glioblastoma Tumor Cells.

This in vitro study aims to evaluate the magnetic hyperthermia (MHT) technique and the best strategy for internalization of magnetic nanoparticles coated with aminosilane (SPIONAmine) in glioblastoma tumor cells. SPIONAmine of 50 and 100 nm were used for specific absorption rate (SAR) analysis, performing the MHT with intensities of 50, 150, and 300 Gauss and frequencies varying between 305 and 557 kHz. The internalization strategy was performed using 100, 200, and 300 µgFe/mL of SPIONAmine, with or without Poly-L-Lysine (PLL) and filter, and with or without static or dynamic magnet field. The cell viability was evaluated after determination of MHT best condition of SPIONAmine internalization. The maximum SAR values of SPIONAmine (50 nm) and SPIONAmine (100 nm) identified were 184.41 W/g and 337.83 W/g, respectively, using a frequency of 557 kHz and intensity of 300 Gauss (≈23.93 kA/m). The best internalization strategy was 100 µgFe/mL of SPIONAmine (100 nm) using PLL with filter and dynamic magnet field, submitted to MHT for 40 min at 44 °C. This condition displayed 70.0% decreased in cell viability by flow cytometry and 68.1% by BLI. We can conclude that our study is promising as an antitumor treatment, based on intra- and extracellular MHT effects. The optimization of the nanoparticles internalization process associated with their magnetic characteristics potentiates the extracellular acute and late intracellular effect of MHT achieving greater efficiency in the therapeutic process.

Multimodal Tracking of Hematopoietic Stem Cells from Young and Old Mice Labeled with Magnetic-Fluorescent Nanoparticles and Their Grafting by Bioluminescence in a Bone Marrow Transplant Model.

This study proposes an innovative way to evaluate the homing and tracking of hematopoietic stem cells from young and old mice labeled with SPIONNIRF-Rh conjugated with two types of fluorophores (NIRF and Rhodamine), and their grafting by bioluminescence (BLI) in a bone marrow transplant (BMT) model. In an in vitro study, we isolated bone marrow mononuclear cells (BM-MNC) from young and old mice, and analyzed the physical-chemical characteristics of SPIONNIRF-Rh, their internalization, cell viability, and the iron quantification by NIRF, ICP-MS, and MRI. The in vivo study was performed in a BMT model to evaluate the homing, tracking, and grafting of young and old BM-MNC labeled with SPIONNIRF-Rh by NIRF and BLI, as well as the hematological reconstitution for 120 days. 5FU influenced the number of cells isolated mainly in young cells. SPIONNIRF-Rh had adequate characteristics for efficient internalization into BM-MNC. The iron load quantification by NIRF, ICP-MS, and MRI was in the order of 104 SPIONNIRF-Rh/BM-MNC. In the in vivo study, the acute NIRF evaluation showed higher signal intensity in the spinal cord and abdominal region, and the BLI evaluation allowed follow-up (11-120 days), achieving a peak of intensity at 30 days, which remained stable around 108 photons/s until the end. The hematologic evaluation showed similar behavior until 30 days and the histological results confirm that iron is present in almost all tissue evaluated. Our results on BM-MNC homing and tracking in the BMT model did not show a difference in migration or grafting of cells from young or old mice, with the hemogram analysis trending to differentiation towards the myeloid lineage in mice that received cells from old animals. The cell homing by NIRF and long term cell follow-up by BLI highlighted the relevance of the multimodal nanoparticles and combined techniques for evaluation.

Current Clinical Trials Protocols and the Global Effort for Immunization against SARS-CoV-2.

Coronavirus disease 2019 (COVID-19) is the biggest health challenge of the 21st century, affecting millions of people globally. The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has ignited an unprecedented effort from the scientific community in the development of new vaccines on different platforms due to the absence of a broad and effective treatment for COVID-19 or prevention strategy for SARS-CoV-2 dissemination. Based on 50 current studies selected from the main clinical trial databases, this systematic review summarizes the global race for vaccine development against COVID-19. For each study, the main intervention characteristics, the design used, and the local or global center partnerships created are highlighted. Most vaccine developments have taken place in Asia, using a viral vector method. Two purified inactivated SARS-CoV-2 vaccine candidates, an mRNA-based vaccine mRNA1273, and the chimpanzee adenoviral vaccine ChAdOx1 are currently in phase III clinical trials in the respective countries Brazil, the United Arab Emirates, the USA, and the United Kingdom. These vaccines are being developed based on a quickly formed network of collaboration.

Noninvasive Tracking of Hematopoietic Stem Cells in a Bone Marrow Transplant Model.

The hematopoietic stem cell engraftment depends on adequate cell numbers, their homing, and the subsequent short and long-term engraftment of these cells in the niche. We performed a systematic review of the methods employed to track hematopoietic reconstitution using molecular imaging. We searched articles indexed, published prior to January 2020, in PubMed, Cochrane, and Scopus with the following keyword sequences: (Hematopoietic Stem Cell OR Hematopoietic Progenitor Cell) AND (Tracking OR Homing) AND (Transplantation). Of 2191 articles identified, only 21 articles were included in this review, after screening and eligibility assessment. The cell source was in the majority of bone marrow from mice (43%), followed by the umbilical cord from humans (33%). The labeling agent had the follow distribution between the selected studies: 14% nanoparticle, 29% radioisotope, 19% fluorophore, 19% luciferase, and 19% animal transgenic. The type of graft used in the studies was 57% allogeneic, 38% xenogeneic, and 5% autologous, being the HSC receptor: 57% mice, 9% rat, 19% fish, 5% for dog, porcine and salamander. The imaging technique used in the HSC tracking had the following distribution between studies: Positron emission tomography/single-photon emission computed tomography 29%, bioluminescence 33%, fluorescence 19%, magnetic resonance imaging 14%, and near-infrared fluorescence imaging 5%. The efficiency of the graft was evaluated in 61% of the selected studies, and before one month of implantation, the cell renewal was very low (less than 20%), but after three months, the efficiency was more than 50%, mainly in the allogeneic graft. In conclusion, our review showed an increase in using noninvasive imaging techniques in HSC tracking using the bone marrow transplant model. However, successful transplantation depends on the formation of engraftment, and the functionality of cells after the graft, aspects that are poorly explored and that have high relevance for clinical analysis.

Excessive cholecalciferol supplementation increases kidney dysfunction associated with intrarenal artery calcification in obese insulin-resistant mice.

Diabetes mellitus accelerates vascular calcification (VC) and increases the risk of end-stage renal disease (ESRD). Nevertheless, the impact of VC in renal disease progression in type 2 diabetes mellitus (T2DM) is poorly understood. We addressed the effect of VC and mechanisms involved in renal dysfunction in a murine model of insulin resistance and obesity (ob/ob), comparing with their healthy littermates (C57BL/6). We analyzed VC and renal function in both mouse strains after challenging them with Vitamin D3 (VitD3). Although VitD3 similarly increased serum calcium and induced bone disease in both strains, 24-hour urine volume and creatinine pronouncedly decreased only in ob/ob mice. Moreover, ob/ob increased urinary albumin/creatinine ratio (ACR), indicating kidney dysfunction. In parallel, ob/ob developed extensive intrarenal VC after VitD3. Coincidently with increased intrarenal vascular mineralization, our results demonstrated that Bone Morphogenetic Protein-2 (BMP-2) was highly expressed in these arteries exclusively in ob/ob. These data depict a greater susceptibility of ob/ob mice to develop renal disease after VitD3 in comparison to paired C57BL/6. In conclusion, this study unfolds novel mechanisms of progressive renal dysfunction in diabetes mellitus (DM) after VitD3 in vivo associated with increased intrarenal VC and highlights possible harmful effects of long-term supplementation of VitD3 in this population.

Poly(3-hydroxi-butyrate-co-3-hydroxy-valerate) (PHB-HV) microparticles loaded with holmium acetylacetonate as potential contrast agents for magnetic resonance images.

INTRODUCTION: Biodegradable polymers that contain radioactive isotopes such as Holmium 166 have potential applications as beta particle emitters in tumor tissues. Also, Ho(III) is paramagnetic, which makes it suitable as a contrast agent for magnetic resonance (MR) images. METHODS: Holmium acetylacetonate (Ho(acac)3) loaded poly(3-hydroxy-butyrate-co-3-hydroxy-valerate) microspheres, with 5% or 8% of 3-hydroxy-valerate (HV), were prepared by emulsification/evaporation process within 20-53 µm size. Microspheres characterization was done using scanning electron microscopy, energy-dispersive X-ray, and infrared spectroscopies. The release of holmium(III) in sodium phosphate buffer (pH 7.4) was followed for 9 days with inductively coupled plasma. Finally, T2 and T2* magnetic resonance images (MRI) were acquired and compared with the MRI of the inclusion complex of holmium acetylacetonate in some ß-cyclodextrins. RESULTS: Holmium acetylacetonate loading, evaluated by thermogravimetry, was up to 20 times higher for copolymer with 5% of HV. It was shown that microspheres loaded with Ho(acac)3 exhibited an accumulation of Ho(III) on their surfaces but were stable over time, as no expressive release of holmium(III) was detected in 9-day exposition to sodium phosphate buffer. Holmium acetylacetonate in both microspheres or inclusion complexes was very efficient in obtaining T2 and T2* weighted images in magnetic resonance, thus, might be used as contrast agents. CONCLUSION: This is the first description of the use of inclusion complexes of holmium acetylacetonate in biodegradable polymers as contrast agents. New investigations are underway to evaluate the resistance of PHB-HV polymer microparticles to nuclear activation to assess their potential for use as radiopharmaceuticals for the treatment of liver cancer.

Evaluation of temperature induction in focal ischemic thermocoagulation model.

The thermocoagulation model, which consists of focal cerebral ischemia with craniectomy, is helpful in studying permanent ischemic brain lesions and has good reproducibility and low mortality. This study analyzed the best conditions for inducing a focal ischemic lesion by thermocoagulation. We investigated parameters such as temperature and thermal dissipation in the brain tissue during induction and analyzed real-time blood perfusion, histological changes, magnetic resonance imaging (MRI), and motor behavior in a permanent ischemic stroke model. We used three-month-old male Wistar rats, weighing 300-350 g. In the first experiment, the animals were divided into four groups (n = 5 each): one sham surgery group and three ischemic lesion groups having thermocoagulation induction (TCI) temperatures of 200°C, 300°C, and 400°C, respectively, with blood perfusion (basal and 30 min after TCI) and 2,3,5-Triphenyl-tetrazolium chloride (TTC) evaluation at 2 h after TCI. In the second experiment, five groups (n = 5 each) were analyzed by MRI (basal and 24 h after TCI) and behavioral tests (basal and seven days after TCI) with the control group added for the surgical effects. The MRI and TTC analyses revealed that ischemic brain lesions expressively evolved, especially at TCI temperatures of 300°C and 400°C, and significant motor deficits were observed as the animals showed a decrease frequency of movement and an asymmetric pattern. We conclude that a TCI temperature of 400°C causes permanent ischemic stroke and motor deficit.

Getting into the brain: liposome-based strategies for effective drug delivery across the blood-brain barrier.

This review summarizes articles that have been reported in literature on liposome-based strategies for effective drug delivery across the blood-brain barrier. Due to their unique physicochemical characteristics, liposomes have been widely investigated for their application in drug delivery and in vivo bioimaging for the treatment and/or diagnosis of neurological diseases, such as Alzheimer's, Parkinson's, stroke, and glioma. Several strategies have been used to deliver drug and/or imaging agents to the brain. Covalent ligation of such macromolecules as peptides, antibodies, and RNA aptamers is an effective method for receptor-targeting liposomes, which allows their blood-brain barrier penetration and/or the delivery of their therapeutic molecule specifically to the disease site. Additionally, methods have been employed for the development of liposomes that can respond to external stimuli. It can be concluded that the development of liposomes for brain delivery is still in its infancy, although these systems have the potential to revolutionize the ways in which medicine is administered.