Down-regulation of miR-199b associated with imatinib drug resistance in 9q34.1 deleted BCR/ABL positive CML patients

Chronic myeloid leukemia (CML) occurs due to t(9,22) (q34;q11) and molecularly BCR/ABL gene fusion. About 15–18% Philadelphia positive CML patients have gene deletions around the translocation breakpoints on 9q34.1. The microRNAs (miRNAs), namely miR-219-2 and miR-199b, centromeric to the ABL1 gene are frequently lost in CML patients. We have designed a study to determine miR-219-2 and miR-199b expression levels which would help to understand the prognosis of imatinib therapy. A total of 150 CML patients were analyzed to identify 9q deletion. Fluorescent in-situ hybridization (FISH) was performed using BCR/ABL dual color, dual fusion probe to study the signal pattern and BAC probes for miR-199b and miR-219-2 (RP11-339B21 and RP11-395P17) to study the miRNA deletions. The expression level of miRNA was analyzed by real-time polymerase chain reaction (RT-PCR). FISH analysis revealed 9q34.1 deletion in 34 (23%) CML patients. The deletions were not detected using BAC probes for miRNAs in 9q deleted patients. The expression analysis showed down-regulation of miR-199b and miR-219-2 in the 9q deleted patients (34 CML) as compared to a pool of patients without deletion. However, miR-199b (9q34.11) was significantly (p = 0.001) down-regulated compared to miR-219-2. The follow-up study showed that the miR-199b was found to be strongly associated with imatinib resistance, as 44.11% patients showed resistance to imatinib therapy. Hence, the deletion in 9q34.1 region (ABL) plays an important role in disease pathogenesis. Eventually, miRNAs can provide new therapeutic strategies and can be used as a prognostic indicator.     

Alkyloxy carbonyl modified hexapeptides as a high affinity compounds for Wnt5A protein in the treatment of psoriasis

Psoriasis is one of the most prevalent chronic inflammatory diseases of the skin. The Wnt pathways have been documented to play essential role in stem cell self-renewal and keratinocyte differentiation in the skin. Antagonizing the Wnt5a protein would emerge as a novel therapeutics in psoriasis treatment. In this view, we have developed and characterized series of compounds by attaching varied tertiary alkyloxy carbonyl groups at the N-terminal end of the hexapeptide (Met-Asp-Gly-Cys-Glu-Leu) bestowed to inhibit Wnt/Ca2+ signaling in psoriasis. Hexapeptide compound with 1,1-diphenylethoxy carbonyl group attached to N-terminal end of hexapeptide demonstrated highest binding affinity amongst all the evaluated compounds. The compound identified in the study can be subjected further for in vitro and in vivo studies for ADMET properties.     

The ROS‐induced cytotoxicity of ascorbate is attenuated by hypoxia and HIF‐1alpha in the NCI60 cancer cell lines

Intravenous application of high‐dose ascorbate is used in complementary palliative medicine to treat cancer patients. Pharmacological doses of ascorbate in the mM range induce cytotoxicity in cancer cells mediated by reactive oxygen species (ROS), namely hydrogen peroxide and ascorbyl radicals. However, little is known about intrinsic or extrinsic factors modulating this ascorbate‐mediated cytotoxicity. Under normoxia and hypoxia, ascorbate IC₅₀ values were determined on the NCI60 cancer cells. The cell cycle, the influence of cobalt chloride‐induced hypoxia‐inducible factor‐1α (HIF‐1α) and the glucose transporter 1 (GLUT‐1) expression (a pro‐survival HIF‐1α‐downstream‐target) were analysed after ascorbate exposure under normoxic and hypoxic conditions. The amount of ascorbyl radicals increased with rising serum concentrations. Hypoxia (0.1% O₂) globally increased the IC₅₀ of ascorbate in the 60 cancer cell lines from 4.5 ± 3.6 mM to 10.1 ± 5.9 mM (2.2‐fold increase, P < 0.001, Mann–Whitney t‐test), thus inducing cellular resistance towards ascorbate. This ascorbate resistance depended on HIF‐1α‐signalling, but did not correlate with cell line‐specific expression of the ascorbate transporter GLUT‐1. However, under normoxic and hypoxic conditions, ascorbate treatment at the individual IC₅₀ reduced the expression of GLUT‐1 in the cancer cells. Our data show a ROS‐induced, HIF‐1α‐ and O₂‐dependent cytotoxicity of ascorbate on 60 different cancer cells. This suggests that for clinical application, cancer patients should additionally be oxygenized to increase the cytotoxic efficacy of ascorbate.     

Structural biology of presenilin 1 complexes

The presenilin genes were first identified as the site of missense mutations causing early onset autosomal dominant familial Alzheimer's disease. Subsequent work has shown that the presenilin proteins are the catalytic subunits of a hetero-tetrameric complex containing APH1, nicastrin and PEN-2. This complex (variously termed presenilin complex or gamma-secretase complex) performs an unusual type of proteolysis in which the transmembrane domains of Type I proteins are cleaved within the hydrophobic compartment of the membrane. This review describes some of the molecular and structural biology of this unusual enzyme complex. The presenilin complex is a bilobed structure. The head domain contains the ectodomain of nicastrin. The base domain contains a central cavity with a lateral cleft that likely provides the route for access of the substrate to the catalytic cavity within the centre of the base domain. There are reciprocal allosteric interactions between various sites in the complex that affect its function. For instance, binding of Compound E, a peptidomimetic inhibitor to the PS1 N-terminus, induces significant conformational changes that reduces substrate binding at the initial substrate docking site, and thus inhibits substrate cleavage. However, there is a reciprocal allosteric interaction between these sites such that prior binding of the substrate to the initial docking site paradoxically increases the binding of the Compound E peptidomimetic inhibitor. Such reciprocal interactions are likely to form the basis of a gating mechanism that underlies access of substrate to the catalytic site. An increasingly detailed understanding of the structural biology of the presenilin complex is an essential step towards rational design of substrate- and/or cleavage site-specific modulators of presenilin complex function.     

Targeted Deletion of Growth Hormone (GH) Receptor in Macrophage Reveals Novel Osteopontin-mediated Effects of GH on Glucose Homeostasis and Insulin Sensitivity in Diet-induced Obesity

We investigated GH action on macrophage (MΦ) by creating a MΦ-specific GH receptor-null mouse model (MacGHR KO). On a normal diet (10% fat), MacGHR KO and littermate controls exhibited similar growth profiles and glucose excursions on intraperitoneal glucose (ipGTT) and insulin tolerance (ITT) tests. However, when challenged with high fat diet (HFD, 45% fat) for 18 weeks, MacGHR KO mice exhibited impaired ipGTT and ITT compared with controls. In MacGHR KO, adipose-tissue (AT) MΦ abundance was increased with skewing toward M1 polarization. Expression of pro-inflammatory cytokines (IL1β, TNF-α, IL6, and osteopontin (OPN)) were increased in MacGHR KO AT stromal vascular fraction (SVF). In MacGHR KO AT, crown-like-structures were increased with decreased insulin-dependent Akt phosphorylation. The abundance of phosphorylated NF-κB and of OPN was increased in SVF and bone-marrow-derived MΦ in MacGHR KO. GH, acting via an NF-κB site in the distal OPN promoter, inhibited the OPN promoter. Thus in diet-induced obesity (DIO), lack of GH action on the MΦ exerts an unexpected deleterious effect on glucose homeostasis by accentuating AT inflammation and NF-κB-dependent activation of OPN expression. These novel results in mice support the possibility that administration of GH could have salutary effects on DIO-associated chronic inflammation and insulin resistance in humans.     

Six months clinical outcome comparison between quadripolar and bipolar left ventricular leads in cardiac resynchronization therapy: A prospective,non-randomized,single-centre observational study

Background and objectivesQuadripolar left ventricular (LV) leads in cardiac resynchronization therapy (CRT) offer multi-vector pacing with different pacing configurations and hence enabling LV pacing at most suitable site with better lead stability. We aim to compare the outcomes between quadripolar and bipolar LV lead in patients receiving CRT.MethodsIn this prospective, non-randomized, single-center observational study, we enrolled 93 patients receiving CRT with bipolar (BiP) (n = 31) and quadripolar (Quad) (n = 62) LV lead between August 2016 to August 2019. Patients were followed for six months, and outcomes were compared with respect to CRT response (defined as ≥5% absolute increase in left ventricle ejection fraction), electrocardiographic, echocardiographic parameters, NYHA functional class improvement, and incidence of LV lead-related complication.ResultsAt the end of six months follow up, CRT with quadripolar lead was associated with better response rate as compared to bipolar pacing (85.48% vs 64.51%; p = 0.03), lesser heart failure (HF) hospitalization events (1.5 vs 2; p = 0.04) and better improvement in HF symptoms (patients with ≥1 NYHA improvement 87.09% vs 67.74%; p = 0.04). There were fewer deaths per 100 patient-year (6.45 vs 9.37; p = 0.04) and more narrowing of QRS duration (Δ12.56 ± 3.11 ms vs Δ7.29 ± 1.87 ms; p = 0.04) with quadripolar lead use. Lead related complications were significantly more with the use of bipolar lead (74.19% vs 41.94%; p = 0.02).ConclusionsOur prospective, non-randomized, single-center observational study reveals that patients receiving CRT with quadripolar leads have a better response to therapy, lesser heart failure hospitalizations, lower all-cause mortality, and fewer lead-related complications, proving its superiority over the bipolar lead.     

Isolation of Bioactive Compounds from Pistacia integerrima with Promising Effects on Reverse Cancer Multidrug Resistance

Russian Journal of Bioorganic Chemistry - Pistacia integerrima grows in some areas of Pakistan and some other south Asian countries such as Afghanistan, India, Nepal, and Myanmar. It is an...     

A framework for incorporating 3D hyperelastic vascular wall models in 1D blood flow simulations

We present a novel framework for investigating the role of vascular structure on arterial haemodynamics in large vessels, with a special focus on the human common carotid artery (CCA). The analysis is carried out by adopting a three-dimensional (3D) derived, fibre-reinforced, hyperelastic structural model, which is coupled with an axisymmetric, reduced order model describing blood flow. The vessel transmural pressure and lumen area are related via a Holzapfel–Ogden type of law, and the residual stresses along the thickness and length of the vessel are also accounted for. After a structural characterization of the adopted hyperelastic model, we investigate the link underlying the vascular wall response and blood-flow dynamics by comparing the proposed framework results against a popular tube law. The comparison shows that the behaviour of the model can be captured by the simpler linear surrogate only if a representative value of compliance is applied. Sobol’s multi-variable sensitivity analysis is then carried out in order to identify the extent to which the structural parameters have an impact on the CCA haemodynamics. In this case, the local pulse wave velocity (PWV) is used as index for representing the arterial transmission capacity of blood pressure waveforms. The sensitivity analysis suggests that some geometrical factors, such as the stress-free inner radius and opening angle, play a major role on the system’s haemodynamics. Subsequently, we quantified the differences in haemodynamic variables obtained from different virtual CCAs, tube laws and flow conditions. Although each artery presents a distinct vascular response, the differences obtained across different flow regimes are not significant. As expected, the linear tube law is unable to accurately capture all the haemodynamic features characterizing the current model. The findings from the sensitivity analysis are further confirmed by investigating the axial stretching effect on the CCA fluid dynamics. This factor does not seem to alter the pressure and flow waveforms. On the contrary, it is shown that, for an axially stretched vessel, the vascular wall exhibits an attenuation in absolute distension and an increase in circumferential stress, corroborating the findings of previous studies. This analysis shows that the new model offers a good balance between computational complexity and physics captured, making it an ideal framework for studies aiming to investigate the profound link between vascular mechanobiology and blood flow.

     

Induction of endogenous glutathione by the chemoprotective agent, 3H-1,2-dithiole-3-thione, in human neuroblastoma SH-SY5Y cells affords protection against peroxynitrite-induced cytotoxicity

Substantial evidence suggests that peroxynitrite generated from the bi-radical reaction of nitric oxide and superoxide is critically involved in the pathogenesis of neurodegenerative disorders, such as Parkinson's disease. Reaction with sulfhydryl (SH)-containing molecules has been proposed to be a major detoxification pathway of peroxynitrite in biological systems. This study was undertaken to determine if chemically elevated intracellular reduced glutathione (GSH), a major SH-containing biomolecule, affords protection against peroxynitrite-mediated toxicity in cultured neuronal cells. Incubation of human neuroblastoma SH-SY5Y cells with the unique chemoprotectant, 3H-1,2-dithiole-3-thione (D3T), led to a significant elevation of cellular GSH in a concentration-dependent fashion. To examine the protective effects of D3T-induced GSH on peroxynitrite-mediated toxicity, SH-SY5Y cells were pretreated with D3T and then exposed to either the peroxynitrite generator, 3-morpholinosydnonimine (SIN-1), or the authentic peroxynitrite. We observed that D3T-pretreated cells showed a markedly increased resistance to SIN-1- or authentic peroxynitrite-induced cytotoxicity, as assessed by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium reduction assay. Conversely, depletion of cellular GSH by buthionine sulfoximine (BSO) caused a marked potentiation of SIN-1- or authentic peroxynitrite-mediated cytotoxicity. To further demonstrate the causal role for GSH induction in D3T-mediated cytoprotection, SH-SY5Y cells were co-treated with BSO to abolish D3T-induced GSH elevation. Co-treatment of the cells with BSO was found to significantly reverse the protective effects of D3T on SIN-1- or authentic peroxynitrite-elicited cytotoxicity. Taken together, this study demonstrates for the first time that D3T can induce GSH in cultured SH-SY5Y cells, and that the D3T-augmented cellular GSH defense affords a marked protection against peroxynitrite-induced toxicity in cultured human neuronal cells.