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Administration of ex vivo expanded somatic myeloid progenitors has been explored as a way to facilitate a more rapid myeloid recovery and improve overall survival after myeloablation. Recent advances in induced pluripotent stem cell (iPSC) technologies have created alternative platforms for supplying off-the-shelf immunologically compatible myeloid progenitors, including cellular products derived from major histocompatibility complex (MHC) homozygous superdonors, potentially increasing the availability of MHC-matching cells and maximizing the utility of stem cell banking. However, the teratogenic and tumorigenic potential of iPSC-derived progenitor cells and whether they will induce alloreactive antibodies upon transfer remain unclear. We evaluated the safety and efficacy of using CD34+CD45+ hematopoietic progenitors derived from MHC homozygous iPSCs (iHPs) to treat cytopenia after myeloablative hematopoietic stem cell (HSC) transplantation in a Mauritian cynomolgus macaque (MCM) nonhuman primate (NHP) model. We demonstrated that infusion of iHPs was well tolerated and safe, observing no teratomas or tumors in the MCMs up to 1 year after HSC transplantation and iHP infusion. Importantly, the iHPs also did not induce significant levels of alloantibodies in MHC-matched or -mismatched immunocompetent MCMs, even after increasing MHC expression on iHPs with interferon-γ. These results support the feasibility of iHP use in the setting of myeloablation and suggest that iHP products pose a low risk of inducing alloreactive antibodies.

PurposeThis paper aims to bring in augmented reality (AR) into navigation systems to rectify the issues mentioned. This paper proposes an AR enhanced navigation system for location automated teller machine (ATM) counters (AR-ATM) and branches of banks based on user’s choice. Upon selecting the ATM, the navigational path to the destination is drawn from the current location, thereby the user can reach the ATM through the optimal path.Design/methodology/approachTraditional navigation systems require users to map with the real world environment as and when required and also may lead to incorrect path due to minor difference in distance. The traditional navigation systems’ also does not take into consideration the ergonomics and safety of the user.FindingsIn this system, a camera lens is used, which is directed down the street at eye level and the application displays the location of ATMs and bank branches and also provides information about the locations like distance and time through the AR superimposed object.Originality/valueThe application also provides indoor navigation, especially in a multi-storeyed building. Experiments are performed on smartphones that support AR, and the results are promising with no lag in time frame of the real object and virtual object. To determine the factors that regulate the suggested AR tracking mechanism, a quantitative evaluation of the experimental data is also performed. The testing of implemented AR-ATM from the standpoint of end-users is undertaken to evaluate real-time usage comfortability, and the results have been determined to be extremely satisfactory.

Allogeneic hematopoietic stem cell transplants (allo-HSCTs) dramatically reduce HIV reservoirs in antiretroviral therapy (ART) suppressed individuals. However, the mechanism(s) responsible for these post-transplant viral reservoir declines are not fully understood. Therefore, we modeled allo-HSCT in ART-suppressed simian-human immunodeficiency virus (SHIV)-infected Mauritian cynomolgus macaques (MCMs) to illuminate factors contributing to transplant-induced viral reservoir decay. Thus, we infected four MCMs with CCR5-tropic SHIV162P3 and started them on ART 6–16 weeks post-infection (p.i.), maintaining continuous ART during myeloablative conditioning. To prevent graft-versus-host disease (GvHD), we transplanted allogeneic MHC-matched α/β T cell-depleted bone marrow cells and prophylactically treated the MCMs with cyclophosphamide and tacrolimus. The transplants produced ~ 85% whole blood donor chimerism without causing high-grade GvHD. Consequently, three MCMs had undetectable SHIV DNA in their blood post-transplant. However, SHIV-harboring cells persisted in various tissues, with detectable viral DNA in lymph nodes and tissues between 38 and 62 days post-transplant. Further, removing one MCM from ART at 63 days post-transplant resulted in SHIV rapidly rebounding within 7 days of treatment withdrawal. In conclusion, transplanting SHIV-infected MCMs with allogeneic MHC-matched α/β T cell-depleted bone marrow cells prevented high-grade GvHD and decreased SHIV-harboring cells in the blood post-transplant but did not eliminate viral reservoirs in tissues.

Road traffic accidents (RTAs) are a major public health concern causing thousands of injuries and premature deaths each year. The purpose of this study to find out the fatal injuries and correlation of injuries with different epidemiological parameters and to find out the measures for the prevention of their causative factors. The present study was conducted on dead bodies sent for post mortem examination involved in road traffic accidents at mortuary of Nizamabad, Govt. Medical College Hospital (Telangana State) during period from January 2017 to December 2018. During the study period out of 1180 of post mortem examinations 208 (17.62 %) were due to RTA. Our study shows that among these 179 (86.05%) victims were males and 29 (13.95%) were females, major victims involved were Pedestrians 64 (30.76%) followed by occupants of four wheelers 63(30.30%) and motor cyclists 54(25.96%). Maximum victims were died due to head injury 103(49.51%) followed by multiple injuries, chest and abdomen injuries. Keywords: Road traffic accidents, Head injury, Pedestrians, Vehicles.

Cellular therapies combined with genetic engineering technologies have emerged as increasingly powerful tools for immunotherapies of cancers and viral infections including HIV infection. The use of induced pluripotent stem cells (iPSCs) as an unlimited source of blood progenitors and terminally differentiated cells can further expand applicability of cellular immunotherapies by offering “off-the-shelf” therapeutic products to fit specific clinical needs for a broad group of patients. In this review, we discuss current advances in lymphoid and myeloid cell differentiation of iPSCs along with challenges in de novo generation of hematopoietic stem cells (HSCs) and allogeneic iPSC products for “off-the-shelf” immunotherapies.

Adoptive therapies with genetically modified somatic T cells rendered HIV resistant have shown promise for AIDS therapy. A renewable source of HIV resistant human T cells from induced pluripotent stem cells (iPSCs) would further facilitate and broaden the applicability of these therapies. Here, we report successful targeting of the CCR5 locus in iPSCs generated from peripheral blood T cells (T-iPSCs) or fibroblasts (fib-iPSCs) from Mauritian Cynomolgus macaques (MCM), using CRISPR/Cas9 technology. We found that CCR5 editing does not affect pluripotency or hematopoietic and T cell differentiation potentials of fib-iPSCs. However, deletion of CCR5 in T-iPSCs leads to selective loss of their T cell redifferentiation potential without affecting myeloid development. T cells and macrophages produced from CCR5-edited MCM- iPSCs did not support replication of the CCR5-tropic simian immunodeficiency viruses SIVmac239 (T-cell tropic) and SIVmac316 (macrophage-tropic). Overall, these studies provide a platform for further exploration of AIDS therapies based on gene-edited iPSCs in a nonhuman primate preclinical model.

BackgroundAllogeneic hematopoietic stem cell transplants (allo-HSCTs) dramatically reduce HIV reservoirs in antiretroviral therapy (ART) suppressed individuals. However, the mechanism(s) responsible for these post-transplant viral reservoir declines are not fully understood but may include pre-transplant conditioning regimens, ART-mediated protection of donor cells, and graft-versus-host (GvH) responses. Therefore, we modeled allo-HSCT in ART-suppressed simian-human immunodeficiency virus (SHIV)-infected Mauritian cynomolgus macaques (MCMs) to illuminate factors contributing to transplant-induced viral reservoir decay.ResultsWe infected four MCMs with CCR5-tropic SHIV162P3 and started ART 6-16 weeks post-infection (p.i.) to establish robust viral reservoirs. We maintained the MCMs on continuous ART during myeloablative conditioning with total body irradiation (TBI) and while transplanting allogeneic MHC-matched α/β T cell-depleted bone marrow cells. Post-transplant, we prophylactically treated the MCMs with cyclophosphamide and tacrolimus to prevent GvH disease (GvHD). The transplants produced ~85% whole blood donor chimerism without causing high-grade GvHD. Consequently, three MCMs had undetectable SHIV DNA in their peripheral blood mononuclear cells post-transplant. However, SHIV-harboring cells persisted in various tissues. We detected viral DNA in lymph node biopsies and terminal analyses of tissues between 38 and 62 days post-transplant. Further, we removed ART from one MCM at 63 days post-transplant, resulting in viral rebound within seven days and viral loads nearing 1×108SHIV RNA copies/ml of plasma after treatment interruption.ConclusionsOur results indicate that myeloablative conditioning and maintaining ART through the peri-transplant period alone are insufficient for eradicating latent viral reservoirs early after allo-HSCTs. Furthermore, our findings suggest that extended ART and GvH responses may be necessary to substantially deplete viral reservoirs after allo-HSCTs.

Allogeneic hematopoietic stem cell transplantation (alloHSCT) is a potentially curative treatment for hematologic cancers and chronic infections such as human immunodeficiency virus (HIV). Its success in these settings is attributed to the ability of engrafting immune cells to eliminate cancer cells or deplete the HIV reservoir (graft-versus-host effect [GvHE]). However, alloHSCT is commonly associated with graft-versus-host diseases (GvHDs) causing significant morbidity and mortality, thereby requiring development of novel allogeneic HSCT protocols and therapies promoting GvHE without GvHD using physiologically relevant preclinical models. Here we evaluated the outcomes of major histocompatibility complex-matched T-cell receptor α/β-depleted alloHSCT in Mauritian cynomolgus macaques (MCMs). Following T-cell receptor α/β depletion, bone marrow cells were transplanted into major histocompatibility complex-identical MCMs conditioned with total body irradiation. GvHD prophylaxis included sirolimus alone in two animals or tacrolimus with cyclophosphamide in another two animals. Posttransplant chimerism was determined by sequencing diagnostic single-nucleotide polymorphisms to quantify the amounts of donor and recipient cells present in blood. Animals treated posttransplant with sirolimus developed nearly complete chimerism with acute GvHD. In the cyclophosphamide and tacrolimus treatment group, animals developed mixed chimerism without GvHD, with long-term engraftment observed in one animal. None of the animals developed cytomegalovirus infection. These studies indicate the feasibility of alloHSCT engraftment without GvHD in an MHC-identical MCM model following complete myeloablative conditioning and anti-GvHD prophylaxis with posttransplant cyclophosphamide and tacrolimus. Further exploration of this model will provide a platform for elucidating the mechanisms of GvHD and GvHE and for testing novel alloHSCT modalities for HIV infection.

Pluripotent stem cells (PSCs) have the potential to provide homogeneous cell populations of T cells that can be grown at a clinical scale and genetically engineered to meet specific clinical needs. OP9-DLL4, a stromal line ectopically expressing the Notch ligand Delta-like 4 (DLL4) is used to support differentiation of PSCs to T-lymphocytes. This article outlines several protocols related to generation of T cells from human and non-human primate (NHP) PSCs, including initial hematopoietic differentiation of PSC on OP9 feeders or defined conditions, followed by coculture of the OP9-DLL4 cells with the PSC-derived hematopoietic progenitors (HPs), leading to efficient differentiation to T lymphocytes. In addition, we describe a protocol for robust T cell generation from hPSCs conditionally expressing ETS1. The presented protocols provide a platform for T cell production for disease modeling and evaluating their use for immunotherapy in large animal models.

Specification of endothelial cells (ECs) into arterial, venous, and lymphatic cells is a crucial process of vascular development, and expanding our knowledge about EC specification from human pluripotent stem cells (hPSCs) will aid the design of optimal strategies for producing desired types of ECs for therapies. In our prior studies, we revealed that hPSC-derived VE-cadherin(V)(+)CD31(+)CD34(+) ECs are heterogeneous and include at least three major subsets with distinct hemogenic properties: V(+)CD43/235a(−)CD73(−) hemogenic endothelial progenitors (HEPs), V(+)CD43(lo)CD235a(+)73(−) angiogenic hematopoietic progenitors (AHPs), and V(+)CD43/235a(−)73(+) non-HEPs. In this study, using angiogenesis assays, we demonstrated that ECs within these subsets have distinct endothelial colony- and tube-forming properties, proliferative and migratory properties, and endothelial nitric oxide synthase and inflammatory cytokine production potentials. Culture of isolated subsets in arterial, venous, and lymphatic conditions revealed that AHPs are skewed toward lymphatic, HEPs toward arterial, and non-HEPs toward venous differentiation in vitro. These findings suggest that selection and enhancement of production of a particular EC subset may aid in generating desirable EC populations with arterial, venous, or lymphatic properties from hPSCs.

Context: Our previous studies showed that all trans retinoic acid (ATRA) ameliorates alcohol-induced toxicity. Hence, we evaluated the efficacy of ATRA and abstention in the regression of alcohol-induced hepatotoxicity. Materials and methods: After ethanol administration to rats for 90 days, the regression of alcohol-induced toxicity was studied by supplementing ATRA at a dose of 100 μg/kg body weight for 30 days. It was also compared with animals in abstention. Results and discussion: Ethanol administration enhanced oxidative stress, activated HSCs and increased collagen deposition. All these alterations were reversed to a certain extent by ATRA supplementation. Conclusions: ATRA had better efficacy than just abstention in reducing ethanol-induced toxicity. The mechanism might be downregulation of CYP2E1, leading to reduced oxidative stress in the hepatocytes and thus impeding NFκB activation, cytokine production, activation of HSC and resulting in the reduction of inflammation and remodelling of fibrosis by modulating MMP and TIMP.

Bone marrow suppression and associated neutropenia that renders the patient more susceptible to infection is a major complication and limiting factor for current chemotherapy. Strategies aimed at neutrophil reconstitution using transfusions or accelerating neutrophil recovery with G-CSF show a limited effect in reducing the rate of infection. To overcome these limitations, administration of ex vivo expanded somatic hematopoietic progenitors (HPs) has been explored as a way to facilitate a more rapid myeloid recovery and improve overall survival following myeloablation. Recent advances in reprogramming and induced pluripotent stem cell (iPSC) technologies have created alternative platforms for off-the-shelf supply of immunologically compatible HPs, including cellular products derived from MHC homozygous superdonors which can increase the degree of MHC matching and may provide a maximum utility for stem cell banking. In these studies, we developed Mauritian cynomolgus macaque (MCM) model to evaluate the utility and safety of CD34 +CD45 + hematopoietic progenitors derived from iPSCs (iHPs) generated from MHC homozygous animals in the treatment of cytopenia following myeloablative stem cell transplantation. MCM iPSCs were generated from MHC homozygous animals and used to generate iHPs in coculture with OP9. Three groups of MCMs underwent myeloablative total body irradiation (TBI) followed by transplantation of cryopreserved autologous CD34 + HSPCs. Four days after autologous HSPC transplantation, animals received transfusion of 30x10 6/kg cryopreserved iCD34 + cells tagged with eGFP or tdTomato from homozygous MHC-matched iPSCs (group 2) or homozygous MHC mismatched iPSCs (group 3). We have demonstrated that infusion of iHPs is safe and well tolerated. No teratoma or tumor formation was observed in animals one year after infusion of iHPs. Although we detected few iPSC derived hematopoietic cells in the blood within 1-week post-infusion, we did not see any significant differences in blood counts or other peripheral blood parameters between all animal groups. Intriguingly we found footprints of iPSC-derived cells in the colon, lymph node, skin and brain specimens collected 25 days after iHP infusion from one animal. This suggests a possibility of migration and retention within these tissues of iHP and iHP-derived myeloid cells. To assess immunogenicity, MHC homozygous iHPs were infused into immunocompetent MCMs across different MHC barriers. 10x10 6/kg of cryopreserved iCD34+ from homozygous iPSCs were transfused every week for 3 weeks, into animals that were divided into 4 groups namely autologous (group I), MHC matched homozygous (group II), MHC matched heterozygous (group III) and MHC mismatch (group IV). Overall, these studies revealed low immunogenicity of MHC homozygous iHPs in MHC matched homozygous and heterozygous animals, while weak immune response was detected following iHP infusion in some MHC mismatched animals. Disclosures Slukvin: Cynata Therapeutics: Consultancy, Current equity holder in publicly-traded company.

SUMMARY Elucidating the pathways that lead to vasculogenic cells, and being able to identify their progenitors and lineage-restricted cells, is critical to the establishment of human pluripotent stem cell (hPSC) models for vascular diseases and development of vascular therapies. Here, we find that mesoderm-derived pericytes (PCs) and smooth muscle cells (SMCs) originate from a clonal mesenchymal progenitor mesenchymoangioblast (MB). In clonogenic cultures, MBs differentiate into primitive PDGFRβ+ CD271+CD73− mesenchymal progenitors, which give rise to proliferative PCs, SMCs, and mesenchymal stem/stromal cells. MB-derived PCs can be further specified to CD274+ capillary and DLK1+ arteriolar PCs with a proinflammatory and contractile phenotype, respectively. SMC maturation was induced using a MEK inhibitor. Establishing the vasculogenic lineage tree, along with identification of stage- and lineage-specific markers, provides a platform for interrogating the molecular mechanisms that regulate vasculogenic cell specification and diversification and manufacturing well-defined mural cell populations for vascular engineering and cellular therapies from hPSCs., In Brief Kumar et al. find that mesodermal pericytes and smooth muscle cells in human pluripotent stem cell cultures originate from a common endothelial and mesenchymal cell precursor, the mesenchymoangioblast. They show how different lineages of mural cells are specified from mesenchymoangioblasts and define stage- and lineage-specific markers for vasculogenic cells., Graphical Abstract

Therapeutic gene delivery to hematopoietic stem cells (HSCs) holds great potential as a life-saving treatment of monogenic, oncologic, and infectious diseases. However, clinical gene therapy is severely limited by intrinsic HSC resistance to modification with lentiviral vectors (LVs), thus requiring high doses or repeat LV administration to achieve therapeutic gene correction. Here we show that temporary coapplication of the cyclic resveratrol trimer caraphenol A enhances LV gene delivery efficiency to human and nonhuman primate hematopoietic stem and progenitor cells with integrating and nonintegrating LVs. Although significant ex vivo, this effect was most dramatically observed in human lineages derived from HSCs transplanted into immunodeficient mice. We further show that caraphenol A relieves restriction of LV transduction by altering the levels of interferon-induced transmembrane (IFITM) proteins IFITM2 and IFITM3 and their association with late endosomes, thus augmenting LV core endosomal escape. Caraphenol A-mediated IFITM downregulation did not alter the LV integration pattern or bias lineage differentiation. Taken together, these findings compellingly demonstrate that the pharmacologic modification of intrinsic immune restriction factors is a promising and nontoxic approach for improving LV-mediated gene therapy.

Toxicity of the pesticide carbofuran (CF) can be alleviated by curcumin, if not for its poor bioavailability. Hence, we investigated the effect of a bioavailable curcumin-galactomannan complex (CGM) on CF-induced neurotoxicity in rats in comparison to that of unformulated standard curcumin (CS). The CF (5 mg/kg b.wt/day) treatment for 90 days produced chronicity model which were treated with either CS or CGM (100 mg/kg b.wt and 250 mg/kg b.wt/day) for another 30 days. Improvement in CF-induced behaviour was evident in endurance, motor co-ordination and pain response on both CS (p

Human pluripotent stem cells (hPSCs) offer the potential to serve as a versatile and scalable source of T cells for immunotherapies, which could be coupled with genetic engineering technologies to meet specific clinical needs. To improve T cell production from hPSCs, it is essential to identify cell subsets that are highly enriched in T cell progenitors and those stages of development at which NOTCH activation induces the most potent T cells. In this study, we evaluated the efficacy of T cell production from cell populations isolated at different stages of hematopoietic differentiation, including mesoderm, hemogenic endothelium (HE), and multipotent hematopoietic progenitors. We demonstrate that KDRhiCD31− hematovascular mesodermal progenitors (HVMPs) with definitive hematopoietic potential produce the highest numbers of T cells when cultured on OP9-DLL4 as compared with downstream progenitors, including HE and multipotent hematopoietic progenitors. In addition, we found that T cells generated from HVMPs have the capacity to expand for 6–7 wk in vitro, in comparison with T cells generated from HE and hematopoietic progenitors, which could only be expanded for 4–5 wk. Demonstrating the critical need of NOTCH activation at the HVMP stage of hematopoietic development to establish robust T cell production from hPSCs may aid in establishing protocols for the efficient off-the-shelf production and expansion of T cells for treating hematologic malignancies.

This is an original post-print of an article published by Taylor & Francis Group in Carbon Management, 9:5, 459-479 on 31/05/2018, available online:https://www.tandfonline.com/doi/full/10.1080/17583004.2018.1476588

The spectrum of antibacterial activities shown by fatty acid fractions of different microalgae, depending on the unsaturation pattern and chain length are still need to be explored. This study provides information on the fatty acid composition of three microalgae, Chlorella marina, Nannochloropsis occulata and Chaetoceros affinis along with their antibacterial activities against selected clinical and fish pathogens. PUFA content of fatty acid fractions followed the order N. occulata > C. affinis > C. marina. Greater abundance of eicosapentanoic acid found in Nannochloropsis occulata (27.39 ± 0.15%) and Chaetoceros affinis (25.34 ± 0.23%) might be responsible for higher bacterial inhibition shown by these fractions.

Background Damaging effects that chronic ethanol exposure causes to the brain and the neurons are well documented. Ethanol and its toxic metabolites increase the oxidative stress in brain. Chronic exposure to ethanol leads to upregulation of N-methyl D-aspartate receptors (NMDAR) and also activates Kruppel like factor 11 (KLF11) mediated death cascade and thereby neurodegeneration. Objective Ethanol depletes vitamin A stores. But supplementation of vitamin A exacerbates ethanol induced toxicity since alcohol and its metabolites are competitive inhibitors of the enzymes involved in the metabolism of vitamin A. Hence, in this study we investigated the impact of co-administration of ethanol and all trans retinoic acid (ATRA), active metabolite of vitamin A, on ethanol induced alterations to the brain. Materials and methods Male Sprague Dawley rats, adolescent, were grouped as follows and maintained for 90 days. I – Control, II – Ethanol (4 g/kg b.w.), III – ATRA (100 µg/kg b.w.), IV – Ethanol (4 g/kg b.w.), +ATRA (100 µg/kg b.w.). Oxidative stress and the mRNA expression of various receptors for the neurotransmitter involved in glutamergic, serotonergic and gabaergic pathways were studied in the brain homogenate. Results Ethanol treatment was shown to decrease brain weight and it was increased on ATRA treatment. Increase in oxidative stress due to ethanol treatment was also brought down on ATRA administration. Ethanol induced upregulation of NMDAR and KLF11 was also downregulated on ATRA supplementation. The alterations in the levels of neurotransmitters and the expression of their receptors due to ethanol treatment also were ameliorated on ATRA supplementation. Conclusion Our results show that ATRA supplementation mitigates the ethanol induced alterations in the brain by reducing oxidative stress in the brain with concurrent suppression of NMDAR and KLF11 expression leading to enhanced catabolism of neurotransmitters.

Molecular pathogenesis of chronic alcoholism is linked to increased endoplasmic reticulum stress. Ethanol is a competitive inhibitor of vitamin A metabolism and vitamin A supplementation aggravates existing liver problems. Hence, we probed into the impact of supplementation of all trans retinoic acid (ATRA), the active metabolite of vitamin A on ethanol-induced endoplasmic reticulcum stress.Male Sprague-Dawley rats were divided into four groups - I: Control; II: Ethanol; III: ATRA; IV: ATRA + Ethanol. After 90 days the animals were sacrificed to study markers of lipid peroxidation in hepatic microsomal fraction and expression of ER stress proteins and apoptosis in liver.Ethanol caused hepatic hyperlipidemia, enhanced microsomal lipid peroxidation, upregulated expression of unfolded protein response associated proteins and that of apoptosis. Ethanol also led to downregulation of retinoid receptors. ATRA supplementation reversed all these alterations indicating the decrease in ethanol-induced endoplasmic reticulum stress.

This report is a part of the China and India Energy Efficiency report series that emerged from the High Impact Opportunities studies supported by t the UNEP DTU Partnership in China and India. In India, the study was carried outby the Indian Institute of Ahmedabad (IIMA) and Energy Efficiency Services Limited (EESL), New Delhi. The othertwo reports published on India areBest Practice and Success Stories on Energy Efficiency in India, andHigh Impact Opportunities for Energy Efficiency in India. This report,Enhancing Energy efficiency in India: Assessment of sectoral potentials, presents a model based assessment ofthe potential for improvements in energy efficiency in key sectors of the Indian economy. It also identifies a set ofshort, medium and long term High Impact Opportunities for energy efficiency, which in combination with varioustechnological options and policies, have potential to provide required future energy services much more efficiently. The process of identification of HIOs included stakeholders’ consultation and f experts opinions on the potential forscaling up the identified technological and policy options.&nbsp

Risk stratification of asymptomatic patients with a Brugada type 1 ECG pattern remains an unresolved clinical conundrum. In contrast to provocative pharmacological testing in Brugada syndrome, there is limited data on the role of exercise stress testing as a risk stratification modality. The objective of this study was to evaluate the utility of exercise testing in asymptomatic patients with type 1 Brugada pattern to prognosticate major arrhythmic events (MAE) during follow-up.Treadmill exercise testing was conducted for 75 asymptomatic patients with type 1 Brugada pattern and for 88 healthy control subjects. The clinical end point of MAE was defined as the occurrence of sudden cardiac death (SCD) or resuscitated ventricular fibrillation (VF). During a follow-up of 77.9 ± 28.9 months, eight MAE occurred (five VF and three SCD). Multivariate Cox regression analysis showed that the following were independent predictors of MAE in asymptomatic patients with a type 1 Brugada pattern: increase in S wave upslope duration ratio30% at peak exercise (HR 1.35, 95% CI 1.08-10.97, P = 0.023), augmentation of J point elevation in lead aVR2 mm in late recovery (HR 1.88, 95% 1.21-15.67, P = 0.011), and delayed HR recovery (HR 1.14, 95% CI 1.06-18.22, P = 0.042). A high-risk cohort was identified by the final step-wise regression model with good accuracy (specificity = 98.4%, sensitivity = 62.5%) and discriminative power (AUC = 0.93, 95% CI 0.89-0.96, P = 0.002). Kaplan-Meier analysis revealed increasing MAE in subjects with one, two, or three predictors, respectively (log rank P0.001).Exercise testing in asymptomatic patients with type 1 Brugada pattern aids in identification of high-risk patients and provides a unique window of opportunity for early intervention.

The angiopoietin/Tie-2 system has been identified as a key role player in tumor angiogenesis. We investigated whether angiopoietin-2 could be a promising target in human neuroblastoma.Angiopoietin-2 down-regulation by siRNA or shRNA was evaluated in vitro in Kelly cells. Angiopoietin-2 shRNA-transfected Kelly cells were tested in a chorioallantoic membrane (CAM) assay to evaluate tumor growth and microvessel density. The effects of L1-10, a peptide-Fc fusion molecule blocking angiopoietin-2/Tie-2 interaction, administered 3 times/week were assessed in a murine neuroblastoma xenograft model.Angiopoietin-2 down-regulation by siRNA or shRNA in Kelly cells inhibited cell proliferation and migration. In vivo growth and microvessel density of angiopoietin-2 shRNA-transfected Kelly cells in the CAM assay were reduced. Therapy of advanced tumors with L1-10 did not stop tumor progression. However, starting L1-10 treatment at the same time as neuroblastoma cell injection significantly inhibited tumor growth (vehicule: 903 ± 160 mm(3); L1-10: 270 ± 152 mm(3) after 26 days; P 0.05). Microvessel density was reduced in both L1-10-treated tumors, whereas expression of angiopoietin-2 and VEGF-A did not change.This first demonstration of beneficial angiopoietin-2 inhibition in neuroblastoma offers an additional approach for future therapy strategies, especially by using L1-10 in the setting of minimal residual disease.

Background Vascular endothelial growth factor (VEGF) is known to play a major role in angiogenesis. A soluble form of Flt-1, a VEGF receptor, is potentially useful as an antagonist of VEGF, and accumulating evidence suggests the applicability of sFlt-1 in tumor suppression. In the present study, we have developed and tested strategies targeted specifically to VEGF for the treatment of ascites formation.Methods As an initial strategy, we produced recombinant sFLT-1 in the baculovirus expression system and used it as a trap to sequester VEGF in the murine ascites carcinoma model. The effect of the treatment on the weight of the animal, cell number, ascites volume and proliferating endothelial cells was studied. The second strategy involved, producing Ehrlich ascites tumor (EAT) cells stably transfected with vectors carrying cDNA encoding truncated form of Flt-1 and using these cells to inhibit ascites tumors in a nude mouse model. Results The sFLT-1 produced by the baculovirus system showed potent antiangiogenic activity as assessed by rat cornea and tube formation assay. sFLT-1 treatment resulted in reduced peritoneal angiogenesis with a concomitant decrease in tumor cell number, volume of ascites, amount of free VEGF and the number of invasive tumor cells as assayed by CD31 staining. EAT cells stably transfected with truncated form of Flt-1 also effectively reduced the tumor burden in nude mice transplanted with these cells, and demonstrated a reduction in ascites formation and peritoneal angiogenesis. Conclusions The inhibition of peritoneal angiogenesis and tumor growth by sequestering VEGF with either sFlt-1 gene expression by recombinant EAT cells or by direct sFLT-1 protein therapy is shown to comprise a potential therapy. Copyright (C) 2009 John Wiley & Sons, Ltd.

1,3,4-Oxadiazoles are an important class of heterocyclic compounds, which play a pivotal role in various pharmaceutical applications. Here, we investigated the antiangiogenic and antiproliferative effects of the derivatives and explored its mechanism of action on EAT cells.The cytotoxic effect of the derivatives on EAT and HEK293 cells was assessed by MTT assay. Effect of the derivatives on ALP activity and proliferation was measured. Swiss albino mice transplanted with EAT cells were used as a model system to study the effect of the derivatives in vivo. Inhibition of angiogenesis in mice peritoneum, CAM and Cornea of the rat were studied. Finally, the effects on VEGF gene expression, HIF-1alpha translocation and cell cycle arrest were determined.The IC50 range for growth inhibition of EAT cells was found to be 140-175 microM. In contrast normal HEK293 cells were resistant to the derivatives at this range. Treatment with derivatives in vivo was demonstrated by the down regulation of VEGF in EAT cells and inhibition of blood vessels formation in mice peritoneum, CAM and cornea of rat, indicating the potent angioinhibitory effect of the derivatives. VEGF promoter-luciferase reporter gene expression analysis showed suppression of VEGF gene expression in vitro. The derivatives proved to be potent antiproliferative agents as shown by FACS analysis and decreased ALP activity. Furthermore, expression of HIF-1alpha was also down regulated by derivatives by repressing its nuclear translocation.Oxadiazole derivatives are strong bioactive compounds with antiangiogenic and antiproliferative potential both in vitro and in vivo. We postulate that diminished HIF-1alpha nuclear presence in oxadiazole treated EAT cells could be responsible for decreased VEGF expression and antiangiogenic effects.

The authors investigate the antiangiogenic and proapoptotic effects of mustard essential oil containing allyl isothiocyanate (AITC) and explore its mechanism of action on Ehrlich ascites tumor (EAT) cells. Swiss albino mice transplanted with EAT cells were used to study the effect of AITC. AITC was effective at a concentration of 10 μm as demonstrated by the inhibition of proliferation of EAT cells when compared with the normal HEK293 cells. It significantly reduced ascites secretion and tumor cell proliferation by about 80% and inhibited vascular endothelial growth factor expression in tumor-bearing mice in vivo. It also reduced vessel sprouting and exhibited potent antiangiogenic activity in the chorioallantoic membrane and cornea of the rat. AITC arrested the growth of EAT cells by inducing apoptosis and effectively arrested cell cycle progression at the G1 phase. The results clearly suggest that AITC inhibits tumor growth by both antiangiogenic and proapoptotic mechanisms.

Nicotine is a parasympathomimetic alkaloid present in tobacco which can induce hyperlipidemia and has a direct effect on neural functions. Statins, competitive inhibitors of 3-hydroxymethyl-3-glutaryl-coenzyme-A reductase, are cholesterol lowering drugs. It has some neuroprotective effects. Hence we analysed the combined effect of nicotine and statin on the learning behaviour of male albino rats. We employed Y-Maze conditional discrimination task. Rats were divided into 4 groups with six rats in each group. (1) Control, (2) Atorvastatin (10mg/kgb.wt), (3) Nicotine (0.6mg/kgb.wt) and (4) Atorvastatin (10mg/kgb.wt)+Nicotine (0.6mg/kgb.wt). After 30days of treatment rats from each group were selected for behavioural study and they were observed for 30days. At the end of the experimental period rats were sacrificed, and brain and liver were dissected out for further biochemical analysis. Nicotine treated group showed least performance in learning in comparison with control, atorvastatin and atorvastatin+nicotine treated groups. Co-administration of atorvastatin and nicotine improved learning behaviour compared to nicotine treated group. Reactive oxygen species level was significantly increased in nicotine group compared to control. The level of neurotransmitter serotonin which has a significant role in learning was found to be decreased in nicotine treated group compared to the control group. Activity of Na(+) K(+) ATPase, Ca(2+) ATPase and glutathione content was significantly reduced in nicotine treated group compared to control. The activity of acetylcholine esterase was significantly increased in the nicotine treated group. Expression studies showed significant decrease in N-methyl D-aspartate receptors and increase in mono amine oxidase-A and mono amine oxidase-B in nicotine treated group and was reversed in atorvastatin + nicotine treated group. It can be concluded that co-administration of nicotine with statin ameliorates the neural functional alterations caused by nicotine to a significant level.

Background aims Existing treatments have limited success in modifying the course of peripheral artery disease, which may eventually lead to limb-threatening ulcers and amputation. Cellular therapies have the potential to provide a new treatment option for this condition, but isolation of cells by conventional means has limitations with respect to reproducibility and scalability. Methods Induced pluripotent stem cells (iPSCs) were differentiated into precursor cells known as mesenchymoangioblasts (MCAs) and subsequently into mesenchymal stromal cells (MSCs). Hindlimb ischemia in mice was created by ligating both the iliac and femoral arteries of one hindlimb. Immediately after surgery, each animal received intramuscular injections of 5 × 10 6 cells or media in the ischemic limb. Toe necrosis was assessed visually, and hindlimb blood flow was measured by laser Doppler using a set region of interest (ROI) and by tracing the entire foot. Myofiber heterogeneity, nuclear centralization, fatty degeneration, fibrosis and capillary angiogenesis in the gastrocnemius muscle were assessed histologically. Results Blood flow in the MCA-derived MSC-treated animals was higher at each day ( P 0.006), and these mice recovered faster than control animals (3.6 vs. 2.5 for set ROI; 7.5 vs. 4.1 foot tracing; slope; P 0.001). There was significantly less myofiber heterogeneity, nuclear centralization, fatty degeneration and fibrosis in MCA-derived MSC-treated animals, indicating less tissue damage. Discussion MCA-derived MSCs improved limb blood flow, reduced necrosis and maintained muscle mass and gross muscle appearance. We conclude that MCA-derived MSCs have a significant and protective effect against ischemic insults.

A major obstacle to the success of gene therapy for hematologic disorders is the inefficiency of lentiviral vector (LV) gene transfer to hematopoietic stem cells (HSCs). Achieving clinically relevant gene delivery levels requires prolonged ex vivo culture of HSCs with cytokines and large LV doses. Rapamycin, PGE2, and other small molecules, have been reported to increase LV transduction of HSCs, however the mechanism of action of these drugs and the basis for LV restriction in HSCs are poorly understood.Here, we report a novel small molecule, β-deliverin, which improves LV gene transfer to HSCs up to 3 fold over DMSO control in vitro. This effect is most pronounced in human adult peripheral blood-derived HSCs, but also observed in human cord-derived HSCs, and in non-human primate bone marrow-derived HSCs. Importantly, treatment with β-deliverin does not significantly affect the viability or expansion of HSCs in culture. Furthermore, for cord-derived HSCs, there was no reduction in the number or type of colony-forming cells. In vivo, β-deliverin treated HSCs engraft in the peripheral blood of NSG mice at levels comparable to control at 16 weeks post-engraftment. Despite an only modest increase in transduction efficiency in β-deliverin-treated cord-derived HSCs transduced in vitro (26% versus 20% for control), the marking frequency for control cells dropped to

Molecular sieves of the MCM-41 type were studied as catalysts for the Friedel-Crafts acylation of 2-methoxynaphthalene in the liquid phase. When MCM-41 in the H + form is used as Bronsted-acid catalyst, acylation with acetic anhydride gives substitution predominantly at the 1-position. Increased reaction temperatures lead to a slight decrease of selectivity to acylation at the 1-position. The regenerability of H-MCM-41 is found to be excellent. For the acylation of 2-methoxynaphthalene with acetyl chloride Zn-MCM-41 was studied as a Lewis-acid catalyst, but both selectivity and regenerability are less favourable than in the case of the H-MCM-41 catalysed acylation of with acid anhydride.

Potter’s Syndrome is a rare congenital malformation which is an atypical physical appearance of the foetus or neonate due to Oligohydramnios experienced in the womb with distinctive facial characteristics. Oligohydramnios is the cause of the various deformities observed in Potter’s Sequence. It is characterised by bilateral renal agenesis, pulmonary hypoplasia, and skeletal defects of the neonate as a direct result of lack of amniotic fluid. Congenital pouch colon (CPC) is an extremely rare variant of anorectal malformation (ARM), in which varying lengths of the colon is replaced by a dilated pouch accompanied by a fistula communicating with the genitourinary tract. Complete Penoscrotal Transposition (CPST) is a rare and unusual malformation in which the scrotum is located cephalic to the penis. It is associated with major and often life threatening malformations. The present case has multiple malformations which include, potter’s syndrome, congenital pouch colon in omphalocele and complete penoscrotal transposition. The aim of this study was to determine the incidence, prevalence of these multiple malformations according to anatomical localization. This complex anomaly is very rare and interesting. Survival is extremely rare. In view of prognosis early diagnosis allows for earlier and less traumatic therapeutic abortion.

Descriptions of Entrapment Neuropathies involving the peripheral nerves are relatively common, especially in Sciatic nerve (SN). Sciatic nerve is the largest nerve in the body. It originates from the sacral plexus from L4-S3 roots in form of two nerve trunks. The Tibial nerve (TN) and Common Peroneal nerve (CPN) are encompassed by single epineural sheath and eventually separates. Variations in the level of bifurcation of the Sciatic nerve are common and being reported by several authors. It is broad and flat at its origin, peripherally it becomes rounded. The bifurcation into its two major trunks TN and CPN may occur anywhere between the sacral plexus and popliteal space. Significant number of variations in the bifurcation, course, relation and distribution of its branches were encountered in sciatic nerve. These variations may cause nerve compressions under other anatomic structures, resulting in non discogenic sciatica. The aim of this study was to provide and define the level of the SN exit, its divisions and its anatomical variation obtained from human cadavers. The differences in the exit of these two branches are important in clarifying clinical etiology.

Induced pluripotent stem cells (iPSCs) have created novel opportunities for the scalable manufacture of blood products for clinical use. Recent advances in hematopoietic differentiation from human pluripotent stem cells have brought the clinical translation of iPSC-derived blood products close to reality and evoke an urgent need for preclinical evaluation of their efficacy, safety, and immunogenicity in large animal models. Due to substantial similarities with humans, outcomes of cellular therapies in nonhuman primate (NHP) models can be readily extrapolated to clinical setting. However, the use of this model is hampered by limited availability of clinically relevant NHP-iPSCs and low efficiency of myeloid and lymphoid differentiation from them. Here, we describe generation of clinically relevant transgene-free iPSCs from different NHP species, including rhesus, Chinese and Mauritian cynomolgus monkeys, and demonstrate that GSK3β inhibition is essential to induce rapid and efficient differentiation of the NHP-iPSCs into multipotential CD34+CD45+CD90+CD38-CD45RA-hematopoietic progenitors in coculture with OP9 bone marrow stromal cells. NHP-iPSC-derived hematopoietic progenitors were capable of differentiating further into mature erythroid cells, megakaryocytes, granulocytes, monocytes/macrophages following exposure to hematopoietic cytokines promoting development corresponding lineages. In addition, iPSC-derived CD34+CD45+ cells generated CD4+CD8+ T lymphoid cells and CD159a+ NK cells when cultured on OP9 cells expressing DLL4. To confirm T cell development, we analyzed the genomic DNA of the hematopoietic cells from OP9-DLL4 cultures for the presence of T cell receptor (TCR) rearrangements. This analysis demonstrated the presence of multiple PCR products of random V-J and D-J rearrangements at the β locus and V-J rearrangements at the γ locus, indicative of a polyclonal T lineage repertoire. To evaluate feasibility of using NHP model for evaluation of safety and clinical efficacy of iPSC-derived blood products, we generated iPSCs from Mauritian cynomolgus macaque and marked them with GFP. GFP-iPSCs were used to generate CD34+CD45+ multipotential hematopoietic progenitors using multiple rounds of differentiation followed by freezing. Subsequently, iPSC donor animal was subjected to nonmyeloablative conditioning with fludarabine (50 mg/m(2)/day for 3 days) together with low-level total body irradiation (2 Gy) and transfused with 30x106/kg autologous iPSC-derived CD34+CD45+ cells. Transfusion of iPSC-derived CD34+CD45+ cells well tolerated and animal recovered without any adverse events. By flow cytometry, GFP-marked cells (up to 1%) were detected in peripheral blood on days 20-30 post-transfusion, predominately in granulocyte, monocyte and platelet compartments. Although, GFP-positive cells mostly disappeared in circulation one month after transfusion, GFP-positive cells were detected in circulation for more than 90 days by genomic PCR. During 6 month follow-up period no signs of myelo- or lympho-proliferative disorder were detected. Overall, these studies lay the foundation for advancing a NHP model for preclinical testing of iPSC-based therapies for blood diseases. Disclosures Slukvin: Cynata: Consultancy, Equity Ownership, Research Funding.

Background Inhibition of tumor angiogenesis is a promising approach for cancer therapy and the Tie-2/angiopoietin pathway appears to play an important role. In the present study, we have developed strategies to explore the therapeutic potential of blocking the Tie-2/angiopoietin pathway by sTie-2. Methods Ehrlich ascites tumor (EAT) cells were stably transfected to overexpress a truncated form of sTie-2. Transfectants were characterized for their in vitro growth behavior and transplanted into nude mice. Furthermore, recombinant sTie-2 produced by the baculovirus expression system was used to sequester angiopoietins in the murine ascites carcinoma model. The effect of sTie-2 treatment alone or in combination with sFLT-1 on the weight of the animal, ascites cell number and volume was studied. Results EAT cells stably transfected with a truncated form of sTie-2 showed no change in cell proliferation in vitro and colony forming in soft agar compared to control cells. However, sTie-2 transfected EAT cells transplanted into nude mice reduced tumor burden and demonstrated a reduction in ascites formation and peritoneal angiogenesis. Recombinant sTie-2 showed angiogenic activity in the tube formation and wound healing assay in vitro. sTie-2 treatment alone or in combination with sFLT-1 in an ascites tumor mouse model resulted in reduced peritoneal angiogenesis, with a concomitant decrease in tumor cell number, volume of ascites and the number of invasive tumor cells, as assayed by CD31 staining. Conclusions The findings of the present study demonstrate an important role for the Tie-2/angiopoietin pathway in the formation of tumor vasculature and suggest that sTie-2 might yield useful anticancer therapy. Copyright © 2010 John Wiley & Sons, Ltd.

The multiple pharmacological actions of unique synthetic compounds are a prerequisite for classifying a drug as highly efficacious, because the multiple pharmacological actions offer the possibility of treating various diseases like cancer. 1,3,4-Oxadiazoles are an important class of heterocyclic compounds with broad spectrum of biological activities. In this study we focused on the ability of these derivatives to induce apoptosis in cultured MCF-7 breast cancer cells. Treatment of MCF-7 cells with varying concentrations of the different derivatives resulted in dose and time dependent sequence of events marked by apoptosis, as shown by loss of cell viability, chromatin condensation, internucleosomal DNA fragmentation and sub G(0) phase accumulation. Furthermore, apoptosis in MCF-7 cell was induced by upregulation of proto-oncoprotein Bax and activation of Caspase-3 activated DNase. Although the derivatives induced apoptosis was associated with Bax protein levels, negligible Bcl-2 reduction was observed. Analysis of the data suggests that the substituted oxadiazole derivatives exert antiproliferative action and growth inhibition on MCF-7 cells through apoptosis induction and that it may have anticancer properties valuable for application in drug products.

Objective: To compare the mean Glycosylated haemoglobin level and lipid profile of children using high dose ICS with that of children using low dose ICS. Study Population: Children between 1-12 years attending asthma clinic using high dose and low dose inhaled corticosteroids. HbA1c levels, Fasting blood sugar and Lipid profile of children in the two groups were measured Results: Children belonging to low dose ICS group were using a mean dose of 206.02 ± 67.13 micrograms per day and high dose group were using a mean dose of 444.61 ± 70.33 micrograms per day. The mean glycosylated haemoglobin level was 6.206 ±1.36% among children using low dose inhaled steroids and 6.013 ± 1.185% among children using high dose inhaled steroids. The mean fasting cholesterol was 180.39 ± 28.66 mg/dl in the low dose ICS group and 179.17 ± 30.21 mg/dl in the high dose ICS group. The mean high density lipoprotein levels were 50.8 3 ±11.05 mg/dl among children using low dose inhaled steroids and 53.37 ± 13.36 mg/dl among children using high dose inhaled steroids. The mean low density lipoprotein levels were 111.78 ± 25.39 mg/dl among children using low dose inhaled steroids and 108.01 ± 30.26 mg/dl among children using high dose inhaled steroids. Conclusions: There was no statistically significant difference between the glycosylated haemoglobin levels, fasting blood glucose levels and fasting lipid profile between children on low dose inhaled corticosteroids and high dose inhaled corticosteroids.

India's contributions to meeting global mean temperature increases of 2 °C or well below 2 °C would require transformational changes in its energy systems. A bottom-up model analyzes alternate futures (reference, intended nationally determined contributions and low-carbon scenarios) assuming equal per-capita cumulative emissions rights from 2011 through 2050. The cumulative CO2 budget for India for low-carbon scenarios during this period is estimated to be around 115 Bt-CO2, as against 165 Bt-CO2 for the reference scenario. To achieve such emission reductions, while maintaining high economic growth and meeting sustainable development goals, the paper projects that a range of endemic transformations are required such as shifting toward cleaner fuels, resource efficient technologies, widespread use of Information and communication technology (ICTs) to balance demand and supply (e.g. smart grids), substituting demand in transport (e.g. work from home), aggressive promotion of renewables, lifestyle changes, and CO2 capture, storage and use. Modeling decarbonization to meet the needs of the increasing population and urbanization faces myriad challenges due to the distributed nature of technologies used to provide various services, involving risks and uncertainties. The paper finally outlines specific opportunities and challenges faced to meet the increased mitigation ambition to limit the warming to 2 °C and below.

India's contributions to meeting global mean temperature increases of 2 °C or well below 2 °C would require transformational changes in its energy systems. A bottom-up model analyzes alternate futures (reference, intended nationally determined contributions and low-carbon scenarios) assuming equal per-capita cumulative emissions rights from 2011 through 2050. The cumulative CO2 budget for India for low-carbon scenarios during this period is estimated to be around 115 Bt-CO2, as against 165 Bt-CO2 for the reference scenario. To achieve such emission reductions, while maintaining high economic growth and meeting sustainable development goals, the paper projects that a range of endemic transformations are required such as shifting toward cleaner fuels, resource efficient technologies, widespread use of Information and communication technology (ICTs) to balance demand and supply (e.g. smart grids), substituting demand in transport (e.g. work from home), aggressive promotion of renewables, lifestyle changes, and CO2 capture, storage and use. Modeling decarbonization to meet the needs of the increasing population and urbanization faces myriad challenges due to the distributed nature of technologies used to provide various services, involving risks and uncertainties. The paper finally outlines specific opportunities and challenges faced to meet the increased mitigation ambition to limit the warming to 2 °C and below.