HSP70/IL-2 treated NK cells effectively cross the blood brain barrier and target tumor cells in a rat model of induced glioblastoma multiforme (GBM)

Natural killer (NK) cell therapy is one of the most promising treatments for Glioblastoma Multiforme (GBM). However, this emerging technology is limited by the availability of sufficient numbers of fully functional cells. Here, we investigated the efficacy of NK cells that were expanded and treated by interleukin-2 (IL-2) and heat shock protein 70 (HSP70), both in vitro and in vivo. Proliferation and cytotoxicity assays were used to assess the functionality of NK cells in vitro, after which treated and naïve NK cells were administrated intracranially and systemically to compare the potential antitumor activities in our in vivo rat GBM models. In vitro assays provided strong evidence of NK cell efficacy against C6 tumor cells. In vivo tracking of NK cells showed efficient homing around and within the tumor site. Furthermore, significant amelioration of the tumor in rats treated with HSP70/Il-2-treated NK cells as compared to those subjected to nontreated NK cells, as confirmed by MRI, proved the efficacy of adoptive NK cell therapy. Moreover, results obtained with systemic injection confirmed migration of activated NK cells over the blood brain barrier and subsequent targeting of GBM tumor cells. Our data suggest that administration of HSP70/Il-2-treated NK cells may be a promising therapeutic approach to be considered in the treatment of GBM. © 2020 by the authors. Licensee MDPI, Basel, Switzerland

Alternate processing of Flt1 transcripts is directed by conserved cis-elements within an intronic region of FLT1 that reciprocally regulates splicing and polyadenylation

The vascular endothelial growth factor receptor, Flt1 is a transmembrane receptor co-expressed with an alternate transcript encoding a secreted form, sFlt1, that functions as a competitive inhibitor of Flt1. Despite shared transcription start sites and upstream regulatory elements, sFlt1 is in far greater excess of Flt1 in the human placenta. Phorbol myristic acid and dimethyloxalylglycine differentially stimulate sFlt1 compared to Flt1 expression in vascular endothelial cells and in cytotrophoblasts. An FLT1 minigene construct containing exon 13, 14 and the intervening region, recapitulates mRNA processing when transfected into COS-7, with chimeric intronic sFlt1 transcripts arising by intronic polyadenylation and other Flt1/sFlt1 transcripts by alternate splicing. Inclusion of exon 15 but not 14 had a modest stimulatory effect on the abundance of sFlt1. The intronic region containing the distal poly(A) signal sequences, when transferred to a heterologous minigene construct, inhibited splicing but only when cloned in sense orientation, consistent with the presence of a directional cis-element. Serial deletional and targeted mutational analysis of cis-elements within intron 13 identified intronic poly(A) signal sequences and adjacent cis-elements as the principal determinants of the relative ratio of intronic sFlt1 and spliced Flt1. We conclude that intronic signals reciprocally regulate splicing and polyadenylation and control sFlt1 expression

Iranian joint registry (iranian national hip and knee arthroplasty registry)

Periodic evaluation and monitoring the health and economic outcome of joint replacement surgery is a common and popular process under the territory of joint registries in many countries. In this article we introduce the methodology used for the foundation of the National Iranian Joint Registry (IJR) with a joint collaboration of the Social Security Organization (SSO) and academic research departments considering the requirements of the Iran's Ministry of Health and Education. ©BY THE ARCHIVES OF BONE AND JOINT SURGERY

Iranian joint registry (iranian national hip and knee arthroplasty registry)

Periodic evaluation and monitoring the health and economic outcome of joint replacement surgery is a common and popular process under the territory of joint registries in many countries. In this article we introduce the methodology used for the foundation of the National Iranian Joint Registry (IJR) with a joint collaboration of the Social Security Organization (SSO) and academic research departments considering the requirements of the Iran's Ministry of Health and Education. ©BY THE ARCHIVES OF BONE AND JOINT SURGERY

Full-length human placental sFlt-1-e15a isoform induces distinct maternal phenotypes of preeclampsia in mice

<div><p>Objective</p><p>Most anti-angiogenic preeclampsia models in rodents utilized the overexpression of a truncated soluble fms-like tyrosine kinase-1 (sFlt-1) not expressed in any species. Other limitations of mouse preeclampsia models included stressful blood pressure measurements and the lack of postpartum monitoring. We aimed to 1) develop a mouse model of preeclampsia by administering the most abundant human placental sFlt-1 isoform (hsFlt-1-e15a) in preeclampsia; 2) determine blood pressures in non-stressed conditions; and 3) develop a survival surgery that enables the collection of fetuses and placentas and postpartum (PP) monitoring.</p><p>Methods</p><p>Pregnancy status of CD-1 mice was evaluated with high-frequency ultrasound on gestational days (GD) 6 and 7. Telemetry catheters were implanted in the carotid artery on GD7, and their positions were verified by ultrasound on GD13. Mice were injected through tail-vein with adenoviruses expressing hsFlt-1-e15a (n = 11) or green fluorescent protein (GFP; n = 9) on GD8/GD11. Placentas and pups were delivered by cesarean section on GD18 allowing PP monitoring. Urine samples were collected with cystocentesis on GD6/GD7, GD13, GD18, and PPD8, and albumin/creatinine ratios were determined. GFP and hsFlt-1-e15a expression profiles were determined by qRT-PCR. Aortic ring assays were performed to assess the effect of hsFlt-1-e15a on endothelia.</p><p>Results</p><p>Ultrasound predicted pregnancy on GD7 in 97% of cases. Cesarean section survival rate was 100%. Mean arterial blood pressure was higher in hsFlt-1-e15a-treated than in GFP-treated mice (∆MAP = 13.2 mmHg, p = 0.00107; GD18). Focal glomerular changes were found in hsFlt-1-e15a -treated mice, which had higher urine albumin/creatinine ratios than controls (109.3±51.7μg/mg vs. 19.3±5.6μg/mg, p = 4.4x10^-2; GD18). Aortic ring assays showed a 46% lesser microvessel outgrowth in hsFlt-1-e15a-treated than in GFP-treated mice (p = 1.2x10^-2). Placental and fetal weights did not differ between the groups. One mouse with liver disease developed early-onset preeclampsia-like symptoms with intrauterine growth restriction (IUGR).</p><p>Conclusions</p><p>A mouse model of late-onset preeclampsia was developed with the overexpression of hsFlt-1-e15a, verifying the <i>in vivo</i> pathologic effects of this primate-specific, predominant placental sFlt-1 isoform. HsFlt-1-e15a induced early-onset preeclampsia-like symptoms associated with IUGR in a mouse with a liver disease. Our findings support that hsFlt-1-e15a is central to the terminal pathway of preeclampsia, and it can induce the full spectrum of symptoms in this obstetrical syndrome.</p></div

Transcriptome and kinome analysis during ethylene-induced growth in Rumex palustris

Complete submergence is one of the most severe abiotic stresses worldwide, both in natural and man-made ecosystems. A limited number of species have evolved mechanisms to deal with these conditions. Rumex palustris, a model for studying plant responses to flooding, manages to outgrow the water, and thus restore contact with the atmosphere, through upward leaf growth (hyponasty) followed by strongly enhanced petiole elongation. These responses are initiated by the gaseous plant hormone ethylene, which accumulates inside plants due to physical entrapment and mimics largely this morphological submergence response. The complexity of ethylene-driven growth in R. palustris upon submergence suggests the involvement of many genes under tight regulation. A comprehensive understanding of the changes in the expression of genes responsive to ethylene is crucial to elucidate the signal transduction networks that lead to hyponastic growth and under-water elongation in this plant. By applying cDNA subtractive hybridisation and cDNA-AFLP (RNA fingerprinting) we identified 119 differentially expressed genes during ethylene-induced hyponastic growth and petiole elongation. From those, 21 have very good homology with Arabidopsis genes. The expression kinetics of these genes were monitored in response to ethylene and submergence using Real Time RTPCR and the putative involvement of candidate genes in (differential) growth during ethylene and submergence treatment was confirmed. Furthermore, we applied whole genome profiling by hybridising mRNA from R. palustris to a CATMA spotted array containing Arabidopsis gene specific tags. This resulted in more than 5000 genes that were differentially regulated in one or both traits. Additionally, the presence of transcription factor binding motifs in the promoters of the differentially expressed genes were examined and demonstrated an over-representation of Abscisic acid responsive elements. Aanalysing the function of putative R. palustris orthologous genes in Arabidopsis during ethylene-induced hyponastic growth revealed that a subset of these genes is likely to control the initial angle of the petiole since, mutations in these genes resulted in constitutive elevated initial petiole angles in Arabidopsis mutant plants compared to the wild type Columbia-0. The others have an effect on either the angle of the petiole or leaf blade angle or both. Mutant analysis revealed a role for PSI-H subunit of photosystem I, glyceraldehyde-3-phosphate dehydrogenase and glutamate synthase in hyponastic growth of Arabidopsis. Furthermore, the effect of auxin response factor ARF8 on hyponastic growth was demonstrated. Reversible protein phosphorylation is a fundamental strategy used by eukaryotes to regulate basic cellular function. Upon a variety of stimuli, kinases phosphorylate downstream kinases usually until the target transcription factor is phosphorylated. Moreover, for many metabolic processes reversible phosphorylation of proteins is a key regulatory mechanism. We examined changes in the kinome profile of R. palustris in response to enhanced levels of ethylene and we discovered differential activity of many kinases. From these experiments, however, we cannot conclude the exact protein kinases involved in ethylene-induced (differential) growth. Our results do show, however, that the kinase regulatory system is strongly regulated during ethylene enriched conditions that induce hyponastic growth and petiole elongation in R. palustris

Transcriptome and kinome analysis during ethylene-induced growth in Rumex palustris

Complete submergence is one of the most severe abiotic stresses worldwide, both in natural and man-made ecosystems. A limited number of species have evolved mechanisms to deal with these conditions. Rumex palustris, a model for studying plant responses to flooding, manages to outgrow the water, and thus restore contact with the atmosphere, through upward leaf growth (hyponasty) followed by strongly enhanced petiole elongation. These responses are initiated by the gaseous plant hormone ethylene, which accumulates inside plants due to physical entrapment and mimics largely this morphological submergence response. The complexity of ethylene-driven growth in R. palustris upon submergence suggests the involvement of many genes under tight regulation. A comprehensive understanding of the changes in the expression of genes responsive to ethylene is crucial to elucidate the signal transduction networks that lead to hyponastic growth and under-water elongation in this plant. By applying cDNA subtractive hybridisation and cDNA-AFLP (RNA fingerprinting) we identified 119 differentially expressed genes during ethylene-induced hyponastic growth and petiole elongation. From those, 21 have very good homology with Arabidopsis genes. The expression kinetics of these genes were monitored in response to ethylene and submergence using Real Time RTPCR and the putative involvement of candidate genes in (differential) growth during ethylene and submergence treatment was confirmed. Furthermore, we applied whole genome profiling by hybridising mRNA from R. palustris to a CATMA spotted array containing Arabidopsis gene specific tags. This resulted in more than 5000 genes that were differentially regulated in one or both traits. Additionally, the presence of transcription factor binding motifs in the promoters of the differentially expressed genes were examined and demonstrated an over-representation of Abscisic acid responsive elements. Aanalysing the function of putative R. palustris orthologous genes in Arabidopsis during ethylene-induced hyponastic growth revealed that a subset of these genes is likely to control the initial angle of the petiole since, mutations in these genes resulted in constitutive elevated initial petiole angles in Arabidopsis mutant plants compared to the wild type Columbia-0. The others have an effect on either the angle of the petiole or leaf blade angle or both. Mutant analysis revealed a role for PSI-H subunit of photosystem I, glyceraldehyde-3-phosphate dehydrogenase and glutamate synthase in hyponastic growth of Arabidopsis. Furthermore, the effect of auxin response factor ARF8 on hyponastic growth was demonstrated. Reversible protein phosphorylation is a fundamental strategy used by eukaryotes to regulate basic cellular function. Upon a variety of stimuli, kinases phosphorylate downstream kinases usually until the target transcription factor is phosphorylated. Moreover, for many metabolic processes reversible phosphorylation of proteins is a key regulatory mechanism. We examined changes in the kinome profile of R. palustris in response to enhanced levels of ethylene and we discovered differential activity of many kinases. From these experiments, however, we cannot conclude the exact protein kinases involved in ethylene-induced (differential) growth. Our results do show, however, that the kinase regulatory system is strongly regulated during ethylene enriched conditions that induce hyponastic growth and petiole elongation in R. palustris

Simulation of enhanced characteristic x rays from a 40-MeV electron beam laser accelerated in plasma

Simulation of x-ray generation from bombardment of various solid targets by quasimonoenergetic electrons is considered. The electron bunches are accelerated in a plasma produced by interaction of 500 mJ, 30 femtosecond laser pulses with a helium gas jet. These relativistic electrons propagate in the ion channel generated in the wake of the laser pulse. A beam of MeV electrons can interact with targets to generate x-ray radiation with keV energy. The MCNP-4C code based on Monte Carlo simulation is employed to compare the production of bremsstrahlung and characteristic x rays between 10 and 100 keV by using two quasi-Maxwellian and quasimonoenergetic energy distributions of electrons. For a specific electron spectrum and a definite sample, the maximum x-ray flux varies with the target thickness. Besides, by increasing the target atomic number, the maximum x-ray flux is increased and shifted towards a higher energy level. It is shown that by using the quasimonoenergetic electron profile, a more intense x ray can be produced relative to the quasi-Maxwellian profile (with the same total energy), representing up to 77% flux enhancement at K_{α} energy