Imaging beta-cell death with a near-infrared probe

Evidence exists for an essential role of beta-cell apoptosis in the pathology of type 1 and type 2 diabetes. Current methods for diabetes-associated apoptosis detection, however, suffer the drawbacks of relying on in situ-based strategies. In this study, we attempted to measure, both in vitro and ex vivo, levels of beta-cell apoptosis in diabetic mice using Cy5.5-labeled annexin V. We used streptozotocin-treated BALB/c mice and NOD mice of different ages as models of type 1 diabetes and db/db mice as a model of type 2 diabetes. With annexin V Cy5.5, we established differences in levels of apoptosis between diabetic and control animals. Intravenously administered annexin V Cy5.5 accumulated in pancreata of diabetic mice but not in nondiabetic controls. Furthermore, its localization was specific to apoptotic events within diabetic islets; its selectivity was supported by transferase-mediated dUTP nick-end labeling staining. Because annexin V defines an early marker of apoptosis and the developed probe is suitable for in vivo administration, it may provide a promising tool for real-time identification in intact animals of the earliest stages of diabetes-associated beta-cell death and for tracing the events that characterize the pathology of the disease.     

Level of Knowledge about Sexuality of People with Mental Disabilities

The paper examines the level of knowledge about sexuality of people with mental disabilities. The research also presents differences resulting from sex and the level of mental retardation. The sample included 24 persons with mental disabilities, who the author of this paper knew very well due to the time spent with them during a summer camp at a club in Vjeverica. Data was collected through direct interviews. Two types of questionnaires on sexuality of people with mental disabilities were used during these interviews. Results indicate that the level of knowledge about sexuality is not sufficient. Specifically low knowledge was shown in the area of the protection of sexual health such as sexually transmitted diseases and methods of protection. A relatively good level of knowledge was shown by the respondents in distinguishing between appropriate and inappropriate ways of sexual behaviour and social understanding of certain situational norms. Differences regarding sex and the level of disability were found. The results indicate the need for additional education on sexuality of both people with mental disabilities and their parents, along with support. Despite the fact that the research was conducted on a small occasional sample, it indicates that further research on this subject is needed.     

In vivo imaging of a diabetogenic CD8+ T cell response during type 1 diabetes progression

Type 1 diabetes is preceded by a long, protracted period of pancreatic islet inflammation by autoreactive lymphocytes. Noninvasive imaging of islet inflammation prior to the onset of hyperglycemia might have diagnostic and therapeutic implications, but this is not currently possible. Here, MRI is used to track, noninvasively, the accumulation diabetogenic CD8+ T-cells during type 1 diabetes progression in nonobese diabetic (NOD) mice. The contrast agent is an MRI probe (MN-NRP-V7) that specifically labels CD8+ T-cells recognizing residues 206-214 of islet-specific glucose-6-phosphatase catalytic subunit related protein (IGRP(206-214)) in the context of the major histocompatibility complex (MHC) class I molecule H-2K(d). This probe consists of superparamagnetic iron oxide nanoparticles (MN) coated with K(d) molecules presenting NRP-V7, a high-avidity mimotope of IGRP(206-214). NOD mice of different ages (5, 8, 15, and 24 weeks) were imaged by MRI before and after a single intravenous injection of MN-NRP-V7 or unmodified MN nanoparticles. MN-NRP-V7 accumulation, as determined by semiquantitative MRI analysis of pancreas-associated T(2) relaxation time, was antigen-specific, age-dependent, and well correlated with the numbers of MN-NRP-V7-labeled CD8+ T-cells recovered from the pancreata of the treated mice. Antigen/MHC-coupled nanoparticles represent a promising new avenue for noninvasive imaging of lymphocyte inflammation in organ-specific autoimmunity and transplantation.     

A Theranostic Small Interfering RNA Nanoprobe Protects Pancreatic Islet Grafts From Adoptively Transferred Immune Rejection

Islet transplantation has recently emerged as an acceptable clinical modality for restoring normoglycemia in patients with type 1 diabetes mellitus (T1DM). The long-term survival and function of islet grafts is compromised by immune rejection–related factors. Downregulation of factors that mediate immune rejection using RNA interference holds promise for improving islet graft resistance to damaging factors after transplantation. Here, we used a dual-purpose therapy/imaging small interfering (si)RNA magnetic nanoparticle (MN) probe that targets β₂ microglobulin (B2M), a key component of the major histocompatibility class I complex (MHC I). In addition to serving as a siRNA carrier, this MN-siB2M probe enables monitoring of graft persistence noninvasively using magnetic resonance imaging (MRI). Human islets labeled with these MNs before transplantation into B2M (null) NOD/scid mice showed significantly improved preservation of graft volume starting at 2 weeks, as determined by longitudinal MRI in an adoptive transfer model (P < 0.05). Furthermore, animals transplanted with MN-siB2M–labeled islets demonstrated a significant delay of up to 23.8 ± 4.8 days in diabetes onset after the adoptive transfer of T cells relative to 6.5 ± 4.5 days in controls. This study demonstrated that our approach could protect pancreatic islet grafts from immune rejection and could potentially be applied to allotransplantation and prevention of the autoimmune recurrence of T1DM in islet transplantation or endogenous islets.Type 1 diabetes mellitus (T1DM) is characterized by the selective and progressive destruction of pancreatic β-cells, leading to insulin dependency and serious complications (1). Pancreatic allotransplantation offers superior glycemic control for T1DM patients and can prevent or even reverse secondary complications (2). However, this procedure is associated with significant mortality and morbidity (3). β-Cell replacement by islet transplantation provides a less invasive alternative T1DM treatment, with reduced antigen load, relative surgical simplicity, and low morbidity (4). Unfortunately, even with the success of the Edmonton immunosuppressive protocol, only 20% of patients remained insulin-independent 3 years after islet transplantation (5). Studies indicate that several factors influence the decrease in islet graft function (6). Among them are immunologic factors that play a critical role because they contribute to innate and adaptive immune rejection (7), recurrence of autoimmunity (8), and toxicity associated with immunosuppressive agents (9,10).The potential for islet grafts to elicit allo- or xenoimmunogenic responses depends on their major histocompatibility complex (MHC) compatibility with the recipient HLA (11). Alloreactivity is significantly reduced In HLA-matched islet transplantation (12); however, limited sources of donor islets and extensive HLA polymorphisms restrict HLA-matched allotransplantation (13), prompting the search for alternative transplant sources.One approach involves the use of xenografts (14), although significant immunologic barriers must be overcome before xenotransplantation becomes a reality in a clinical setting (15). As such, a T cell–mediated immune response occurring within weeks or even days of grafting causes irreversible β-cell damage (16). Major immune contributors to this damage include cytotoxic cluster of differentiation (CD)8⁺ T cells that recognize antigenic peptides in context with MHC class I molecules in allo- and xenografts (17,18). Reduction or downregulation of MHC class I protein expression using RNA interference (RNAi) (19) has shown some success in overcoming the limitations of immune rejection in cell-based therapies. The delivery of short hairpin RNA to HeLa cells resulted in a selective and permanent silencing of MHC class I by up to 90%, even under inflammatory conditions (20). MHC class I knockdown was effective in preventing antibody-mediated cell lysis and CD8⁺ T cell response (20). Lentivirus-mediated silencing of HLA in human 293 cells promoted resistance to killing by alloreactive T-effector cells (21) and showed enhancement in hematopoietic stem cell transplantation (22). Encouraged by these results, we sought to use RNAi to disturb MHC class I expression in human islet cells through the silencing of β₂ microglobulin (B2M), a key component of MHC class I molecules, thereby reducing graft rejection in a model of xenotransplantation.Because a safe and highly efficient delivery method is essential for the eventual clinical application of RNAi in islet grafts, we used magnetic nanoparticles (MNs) as a delivery vehicle for B2M small interfering (si)RNA into islet cells. This approach was selected rather than viral vector nucleotide delivery because it is not undermined by excessive inflammation (23) or the potential for oncogenicity (24). In addition to serving as a siRNA carrier, the MNs enable the monitoring of graft persistence noninvasively using magnetic resonance imaging (MRI) (25,26). Treatment of human islets with these dual-purpose therapeutic/imaging MNs before transplantation resulted in significantly improved survival of grafts and a delay in the onset of hyperglycemia after the adoptive transfer of T cells to animals with newly established xenografts. Our approach provides for protection of transplanted islet grafts and can be potentially extended for various transplantation and cell therapy applications.     

Derivation of whole body bone measures through summation of individual DXA scans.

A major limitation of dual-energy X-ray absorptiometry (DXA) instruments is the size of the scanning region that precludes obtaining accurate data on individuals who are very wide and/or tall. The purpose of this study was to determine if whole body bone mineral values can be determined in individuals too wide and/or too tall for the scanning region using summed measures from multiple DXA scans. Two to four partial scans of the body were combined to predict bone mineral content (BMC) and bone mineral density (BMD) from a complete whole body (WB) scan. The results indicate that compared with the WB scan, there were strong relations and small differences when summing scans to simulate a wide or tall individual for BMC (wide: r = 0.999, mean [M] +/- standard deviation(diff) [SD](diff) = -2.1 +/- 26.2 g, p = 0.62; tall: r = 0.999; M +/- SD(diff) = -5.9 +/- 27.1 g, p = 0.17) or BMD (wide: r = 0.996; M +/- SD(diff) = 0.001 +/- 0.011 g/cm(2), p = 0.54; tall: r = 0.994; M +/- SD(diff) = 0.004 +/- 0.014 g/cm(2), p = 0.10). Notably the differences from the whole body scan were within measurement error. In conclusion, accurate assessment of bone mineral outcomes can be obtained on wide or tall individuals using summed values from multiple DXA scans.     

Robotic system for prostate brachytherapy.

In contemporary brachytherapy procedures, needle placement at the desired target is challenging for a variety of reasons. A robot-assisted brachytherapy system can potentially improve needle placement and seed delivery, resulting in enhanced therapeutic outcome. In this paper we present a robotic system with 16 degrees of freedom (DOF) (9 DOF for the positioning module and 7 DOF for the surgery module) that has been developed and fabricated for prostate brachytherapy. Strategies to reduce needle deflection and target movement were incorporated after extensive experimental validation. Provision for needle motion and force feedback was included in the system to improve robot control and seed delivery. Preliminary experimental results reveal that the prototype system is sufficiently accurate in placing brachytherapy needles.