Glycyrrhizic acid modulates t-BHP induced apoptosis in primary rat hepatocytes

Glycyrrhizic acid (GA) is the main bioactive ingredient of licorice (Glycyrrhiza glabra). The object of this study was to evaluate the protective effects of GA on tert-butyl hydroperoxide (t-BHP) induced oxidative injury leading to apoptosis in cultured primary rat hepatocytes. Throughout the study silymarin was used as positive control. Molecular mechanisms involved in apoptotic pathways induced in hepatocytes by t-BHP at 250 μM were explored in detail. DNA fragmentation, activation of caspases and cytochrome c release were demonstrated. In addition, changes in the mitochondrial membrane potential and ROS generation were detected confirming involvement of mitochondrial pathway. Pre-treatment with GA (4 μg) protected the hepatocytes against t-BHP induced oxidative injury and the results were comparable to the pre-treatment with positive control, i.e. silymarin. The protective potential against cell death was achieved mainly by preventing intracellular GSH depletion, decrease in ROS formation as well as inhibition of mitochondrial membrane depolarization. GA was found to modulate critical end points of oxidative stress induced apoptosis and could be beneficial against liver diseases where oxidative stress is known to play a crucial role.     

Rottlerin inhibits human T cell responses

Rottlerin is a pharmacological inhibitor of protein kinase C (PKC) theta, a novel PKC selectively expressed in T lymphocytes. PKC theta is known to regulate T cell receptor (TCR)/CD28 signalling pathways in T lymphocytes, but the impact of PKC theta inhibition on human T cell responses remains undefined. In this work, we describe the effects of rottlerin on the responses of CD4+ and CD8+ human T lymphocytes upon polyclonal activation. We observed a dose-dependent inhibition of CD4+ and CD8+ T cell proliferation in response to anti-CD3/anti-CD28 antibodies stimulation in the presence of rottlerin. This inhibition was associated with impaired CD25 expression and decreased interleukin (IL)-2 production in activated T cells. In contrast, rottlerin did not alter IL-2-induced T cell proliferation. Furthermore, we demonstrated that rottlerin blocked interferon (IFN) gamma, IL-10 and IL-13 mRNA expression in TCR/CD28 activated CD4+ T cells. These findings place rottlerin as a potent immunosuppressive agent for the development of novel therapies in T cell mediated immune disorders.     

Design and cellular kinetics of dansyl-labeled CADA derivatives with anti-HIV and CD4 receptor down-modulating activity

A new class of anti-retrovirals, cyclotriazadisulfonamide (CADA) and its derivatives, specifically down-regulate CD4, the main receptor of HIV, and prevent HIV infection in vitro. In this work, several CADA derivatives, chemically labeled with a fluorescent dansyl group, were evaluated for their biological features and cellular uptake kinetics. We identified a derivative KKD-016 with antiviral and CD4 down-modulating capabilities similar to those of the parental compound CADA. By using flow cytometry, we demonstrated that the dose-dependent cellular uptake of this derivative correlated with CD4 down-modulation. The uptake and activity of the dansyl-labeled compounds were not dependent on the level of expression of CD4 at the cell surface. Removal of the CADA compounds from the cell culture medium resulted in their release from the cells followed by a complete restoration of CD4 expression. The inability of several fluorescent CADA derivatives to down-modulate CD4 was not associated with their lower cellular uptake and was not reversed by facilitating their cell penetration by a surfactant. These results prove the successful integration of the dansyl fluorophore into the chemical structure of a CD4 down-modulating anti-HIV compound, and show the feasibility of tracking a receptor and its down-modulator simultaneously. These fluorescent CADA analogs with reversible CD4 down-regulating potency can now be applied in further studies on receptor modulation, and in the exploration of their potentials as preventive and therapeutic anti-HIV drugs.     

Multifidi Muscle Characteristics and Physical Function Among Older Adults With and Without Chronic Low Back Pain

Objective

To determine whether multifidi size, intramuscular fat, or both, are associated with self-reported and performance-based physical function in older adults with and without chronic low back pain (LBP).

Pre-transplant Screening for Non-HLA Antibodies: Who should be Tested?

Retrospective studies of angiotensin II type 1 receptor antibodies (AT1R-Ab) and anti-endothelial cell antibodies (AECA) have linked these antibodies to allograft injury. Because rising healthcare costs dictate judicious use of laboratory testing, we sought to define characteristics of kidney transplant recipients who may benefit from screening for non-HLA antibodies. Kidney recipients transplanted between 2011 and 2016 at Johns Hopkins, were evaluated for AT1R-Ab and AECA. Pre-transplant antibody levels were compared to clinical and biopsy indications of graft dysfunction. Biopsies were graded using the Banff' 2009–2013 criteria. AT1R-Ab and AECA were detected using ELISA and endothelial cell crossmatches, respectively. AT1R-Ab levels were higher in patients who were positive for AECAs. Re-transplanted patients (p?<?0.0001), males (p?=?0.008) and those with FSGS (p?=?0.04) and younger (p?=?0.04) at time of transplantation were more likely to be positive for AT1R-Ab prior to transplantation. Recipients who were positive for AT1R-Ab prior to transplantation had increases in serum creatinine within 3?months post-transplantation (p?<?0.0001) and developed abnormal biopsies earlier than did AT1R-Ab negative patients (126?days versus 368?days respectively; p?=?0.02). Defining a clinical protocol to identify and preemptively treat patients at risk for acute rejection with detectable non-HLA antibodies is an important objective for the transplant community.     

Influence of immunosuppressive drugs on the CD30 molecule in kidney transplanted patients

Background

Soluble CD30 (sCD30) is a suggested marker for kidney transplantation outcomes. We investigated whether sCD30 serum levels are influenced by immunosuppression and whether they correlate with findings in protocol biopsies and with CD30 gene expression in peripheral blood mononuclear cells (PBMC).

Impact of persistent and cleared preformed HLA DSA on kidney transplant outcomes

Preformed HLA donor-specific antibodies (DSA) only detected with Luminex have been associated with increased risk of antibody-mediated rejection (ABMR) and graft failure after kidney transplantation (KT). Their evolution after KT may modify this risk. We analyzed postransplant evolution of preformed DSA identified retrospectively and their impact on outcomes of 370 KT performed 2006–2014. Antibodies were monitored prospectively at 1-3-5?years after KT and if any dysfunction. Early acute ABMR was more frequent among patients with preformed DSA class-I or I?+?II than isolated class-II (29.4% vs 4.5%, p?=?0.02). One year post-KT, 20 of 34 patients with functioning KT had persistent DSA. Preformed DSA class-II persisted more frequently than class-I/I?+?II (66.7% vs 33.3%; p?=?0.031). The only risk factor independently associated with persistence was pretransplant MFI. Patients with de novo DSA had the highest risk of ABMR (HR 22.2 [CI 6.1–81.2]). Although recipients with persisting preformed DSA had significantly increased ABMR risk (HR 14.7 [CI 6.5–33.0]), those with cleared preformed DSA also had a higher risk than those without DSA (HR 7.01 [CI 2.2–21.8]).Preformed DSA are a very important risk factor for ABMR and graft loss. Patients who clear preformed DSA still show an increased risk of ABMR and graft loss after KT.     

Root cause analysis of limitations of virtual crossmatch for kidney allocation to highly-sensitized patients

Efficient allocation of deceased donor organs depends upon effective prediction of immunologic compatibility based on donor HLA genotype and recipient alloantibody profile, referred to as virtual crossmatching (VCXM). VCXM has demonstrated utility in predicting compatibility, though there is reduced efficacy for patients highly sensitized against allogeneic HLA antigens. The recently revised deceased donor kidney allocation system (KAS) has increased transplantation for this group, but with an increased burden for histocompatibility testing and organ sharing. Given the limitations of VCXM, we hypothesized that increased organ offers for highly-sensitized patients could result in a concomitant increase in offers rejected due to unexpectedly positive crossmatch. Review of 645 crossmatches performed for deceased donor kidney transplantation at our center did not reveal a significant increase in positive crossmatches following KAS implementation. Positive crossmatches not predicted by VCXM were concentrated among highly-sensitized patients. Root cause analysis of VCXM failures identified technical limitations of anti-HLA antibody testing as the most significant contributor to VCXM error. Contributions of technical limitations including additive/synergistic antibody effects, prozone phenomenon, and antigens not represented in standard testing panels, were evaluated by retrospective testing. These data provide insight into the limitations of VCXM, particularly those affecting allocation of kidneys to highly-sensitized patients.     

A simple approach for enhanced immune response using engineered dendritic cell targeted nanoparticles

In this study, we demonstrate a simple strategy for enhanced immune response using a two-component dendritic cell (DC) targeted antigen delivery system. One component consists of a recombinant bifunctional fusion protein (bfFp) used for DC targeting, whereas, the other component is made of biotinylated PLGA nanoparticles that encapsulate the antigen. The fusion protein (bfFp) made of a truncated core-streptavidin fused to anti-DEC-205 single chain antibody (scFv) was mixed with ovalbumin-loaded biotinylated NPs that were formulated using biotin–PEG (2000)–PLGA, and the combination, bfFp functionalized NPs was used for DC targeted antigen delivery. In vitro DC uptake studies revealed a 2-fold higher receptor-mediated uptake of bfFp functionalized NPs when compared to non-targeted NPs. Immunization of the mice with the bfFp functionalized NPs in conjunction with DC maturation stimulus (anti-CD40 mAb) enhanced OVA-specific IgG and IgG subclass responses. Splenocytes of these mice secreted significantly higher levels of Th1 (IFN-γ and IL-2) cytokines upon ex vivo restimulation with OVA. The promising outcomes of the bfFp functionalized DC targeted system support its use as a versatile vaccine delivery system for the design of monovalent or polyvalent vaccines.