Intracoronary boluses of adenosine and sodium nitroprusside in combination reverses slow/no-reflow during angioplasty: a clinical scenario of ischemic preconditioning

No or slow reflow following percutaneous coronary intervention (PCI), despite the presence of a patent epicardial vessel, is a serious complication resulting in increased morbidity and mortality. In the present study, we have evaluated the combination therapy of adenosine and sodium nitroprusside administered as sequential intracoronary (IC) boluses on no-reflow during PCI. Seventy-five high risk acute coronary syndrome patients who underwent PCI with evidence of initial less than TIMI (thrombolysis in myocardial infarction) III flow or developed deterioration in TIMI flow during the procedure were randomized to prophylactic administration of multiple boluses of IC saline solution, adenosine (12 microg/bolus) or the combination of adenosine (12 microg/bolus) and sodium nitroprusside (50 microg/bolus), sequentially. Assessment of TIMI and the TMP (tissue myocardial perfusion) grade was done and major adverse cardiac events (MACE) were assessed at the end of 6 months. Slow or no-reflow was persistent in 70% patients receiving saline solution, 31% patients receiving adenosine, and 4% patient receiving the combination. IC injection with saline solution did not produce improvement in TIMI flow or TMP grade. IC injection with combination resulted in greater improvement of TIMI flow and TMP grade. The crossover of patients with no-reflow in saline solution group or adenosine with combination treatment was associated with reestablishment of TIMI II in 4 and TIMI III in 20 patients. Our data suggest that combination therapy of adenosine and nitroprusside is safe and provides better improvement in coronary flow and MACE as compared with IC adenosine alone in cases of impaired flow during coronary interventions.     

The prevalence of gene duplications and their ancient origin in <Emphasis Type="Italic">Rhodobacter sphaeroides</Emphasis> 2.4.1

Background  

Rhodobacter sphaeroides 2.4.1 is a metabolically versatile organism that belongs to α-3 subdivision of Proteobacteria. The present study was to identify the extent, history, and role of gene duplications in R. sphaeroides 2.4.1, an organism that possesses two chromosomes.     

Expression of 3-hydroxy-3-methylglutaryl-CoA reductase, p-hydroxybenzoate-m-geranyltransferase and genes of phenylpropanoid pathway exhibits positive correlation with shikonins content in arnebia [Arnebia euchroma (Royle) Johnston]

Background

Formation of site specific genomic double strand breaks (DSBs), induced by the expression of a pair of engineered zinc-finger nucleases (ZFNs), dramatically increases the rates of homologous recombination (HR) between a specific genomic target and a donor plasmid. However, for the safe use of ZFN induced HR in practical applications, possible adverse effects of the technology such as cytotoxicity and genotoxicity need to be well understood. In this work, off-target activity of a pair of ZFNs has been examined by measuring the ratio between HR and illegitimate genomic integration in cells that are growing exponentially, and in cells that have been arrested in the G2/M phase.

Results

A reporter cell line that contained consensus ZFN binding sites in an enhanced green fluorescent protein (EGFP) reporter gene was used to measure ratios between HR and non-homologous integration of a plasmid template. Both in human cells (HEK 293) containing the consensus ZFN binding sites and in cells lacking the ZFN binding sites, a 3.5 fold increase in the level of illegitimate integration was observed upon ZFN expression. Since the reporter gene containing the consensus ZFN target sites was found to be intact in cells where illegitimate integration had occurred, increased rates of illegitimate integration most likely resulted from the formation of off-target genomic DSBs. Additionally, in a fraction of the ZFN treated cells the co-occurrence of both specific HR and illegitimate integration was observed. As a mean to minimize unspecific effects, cell cycle manipulation of the target cells by induction of a transient G2/M cell cycle arrest was shown to stimulate the activity of HR while having little effect on the levels of illegitimate integration, thus resulting in a nearly eight fold increase in the ratio between the two processes.

Conclusions

The demonstration that ZFN expression, in addition to stimulating specific gene targeting by HR, leads to increased rates of illegitimate integration emphasizes the importance of careful characterization of ZFN treated cells. In order to reduce off-target events, reversible cell cycle arrest of the target cells in the G2/M phase is an efficient way for increasing the ratio between specific HR and illegitimate integration.     

Plasma arginine-vasopressin following experimental stroke: effect of osmotherapy.

Neurohumoral responses have been implicated in the pathogenesis of ischemia-evoked cerebral edema. In a well-characterized animal model of ischemic stroke, the present study was undertaken to 1) study the profile of plasma arginine-vasopressin (AVP), and 2) determine whether osmotherapy with mannitol and various concentrations of hypertonic saline (HS) solutions influence plasma AVP levels. Halothane-anesthetized adult male Wistar rats were subjected to 2 h of middle cerebral artery occlusion with the intraluminal filament technique. Plasma AVP levels (means +/- SD) were significantly elevated at 24 h (42 +/- 21 pg/ml), 48 h (50 +/- 28 pg/ml), and 72 h (110 +/- 47 pg/ml), and returned to baseline at 96 h (22 +/- 15 pg/ml) following middle cerebral artery occlusion compared with sham-operated controls (14 +/- 7 pg/ml). Plasma AVP levels at 72 h were significantly attenuated with 7.5% HS (37 +/- 8 pg/ml; 360 +/- 11 osmol/l) compared with 0.9% saline (73 +/- 6; 292 +/- 6 osmol/l), 3% HS (66 +/- 8 pg/ml; 303 +/- 12 osmol/l), or mannitol (74 +/- 9 pg/ml; 313 +/- 14 osmol/l) treatment. HS (7.5%) significantly attenuated water content in the ipsilateral and contralateral hemispheres compared with surgical shams, 0.9% saline, 3% HS, and mannitol treatments. Peak plasma AVP levels were not associated with direct histopathological injury to the anterior hypothalamus. Attenuation of brain water content with 7.5% HS treatment coincides with attenuated serum AVP levels, and we speculate that this may represent one additional mechanism by which osmotherapy attenuates edema associated with ischemic stroke.     

Single-particle cryo-EM reveals conformational variability of the oligomeric VCC β-barrel pore in a lipid bilayer

Vibrio cholerae cytolysin (VCC) is a water-soluble, membrane-damaging, pore-forming toxin (PFT) secreted by pathogenic V. cholerae, which causes eukaryotic cell death by altering the plasma membrane permeability. VCC self-assembles on the cell surface and undergoes a dramatic conformational change from prepore to heptameric pore structure. Over the past few years, several high-resolution structures of detergent-solubilized PFTs have been characterized. However, high-resolution structural characterization of small β-PFTs in a lipid environment is still rare. Therefore, we used single-particle cryo-EM to characterize the structure of the VCC oligomer in large unilamellar vesicles, which is the first atomic-resolution cryo-EM structure of VCC. From our study, we were able to provide the first documented visualization of the rim domain amino acid residues of VCC interacting with lipid membrane. Furthermore, cryo-EM characterization of lipid bilayer–embedded VCC suggests interesting conformational variabilities, especially in the transmembrane channel, which could have a potential impact on the pore architecture and assist us in understanding the pore formation mechanism.     

Naive stem cell blastocyst model captures human embryo lineage segregation

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Surgical Retrieval,Isolation and In vitro Expansion of Human Anterior Cruciate Ligament-derived Cells for Tissue Engineering Applications

Injury to the ACL is a commonly encountered problem in active individuals. Even partial tears of this intra-articular knee ligament lead to biomechanical deficiencies that impair function and stability. Current options for the treatment of partial ACL tears range from nonoperative, conservative management to multiple surgical options, such as: thermal modification, single-bundle repair, complete reconstruction, and reconstruction of the damaged portion of the native ligament. Few studies, if any, have demonstrated any single method for management to be consistently superior, and in many cases patients continue to demonstrate persistent instability and other comorbidities.The goal of this study is to identify a potential cell source for utilization in the development of a tissue engineered patch that could be implemented in the repair of a partially torn ACL. A novel protocol was developed for the expansion of cells derived from patients undergoing ACL reconstruction. To isolate the cells, minced hACL tissue obtained during ACL reconstruction was digested in a Collagenase solution. Expansion was performed using DMEM/F12 medium supplemented with 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin (P/S). The cells were then stored at -80 ºC or in liquid nitrogen in a freezing medium consisting of DMSO, FBS and the expansion medium. After thawing, the hACL derived cells were then seeded onto a tissue engineered scaffold, PLAGA (Poly lactic-co-glycolic acid) and control Tissue culture polystyrene (TCPS). After 7 days, SEM was performed to compare cellular adhesion to the PLAGA versus the control TCPS. Cellular morphology was evaluated using immunofluorescence staining. SEM (Scanning Electron Microscope) micrographs demonstrated that cells grew and adhered on both PLAGA and TCPS surfaces and were confluent over the entire surfaces by day 7. Immunofluorescence staining showed normal, non-stressed morphological patterns on both surfaces. This technique is promising for applications in ACL regeneration and reconstruction.     

Gender-Affirming Surgery: Perioperative Medical Care

ObjectiveGender-affirming surgeries are becoming increasingly common in the United States. For many transgender and gender-diverse patients, gender-affirming surgery is a critical aspect of their overall health and wellness and has a significant impact on their social functioning. Although often the role of the primary care provider, endocrinology specialists may also need to counsel their patients and collaborate with surgical teams.MethodsThis narrative review provides an overview of the preoperative assessment and perioperative management for the most common gender-affirming surgeries.ResultsPreoperative assessment prior to gender-affirming surgeries can resemble assessment for other surgeries.ConclusionIn the absence of data specific to gender-affirming surgeries, risk stratification and amelioration must use surrogate data from other populations.