Pericytes and their potential in regenerative medicine across species.

The discovery that pericytes are in vivo counterparts of Mesenchymal Stem/Stromal Cells (MSCs) has placed these perivascular cells in the research spotlight, bringing up hope for a well-characterized cell source for clinical applications, alternative to poorly defined, heterogeneous MSCs preparations currently in use. Native pericytes express typical MSC markers and, after isolation by fluorescence-activated cell sorting, display an MSC phenotype in culture. These features have been demonstrated in different species, including humans and horses, the main targets of regenerative treatments. Significant clinical potential of pericytes has been shown by transplantation of human cells into rodent models of tissue injury, and it is hoped that future studies will demonstrate clinical potential in veterinary species. Here, we provide an overview of the current knowledge on pericytes across different species including humans, companion and large animal models, in relation to their identification in different body tissues, methodology for prospective isolation, characterization, and potential for tissue regeneration. © 2017 International Society for Advancement of Cytometry.

Catalytic Enolate Arylation with 3-Bromoindoles Allows the Formation of ß-Carbolines.

Synthesis of substituted ß-carbolines was accomplished by utilizing the catalytic enolate arylation reaction of ketones in conjunction with several 3-bromoindole derivatives. Quenching of the arylation reaction in situ with an electrophile allowed ready incorporation of functionality at the carboline C-4 position in an efficient one-pot protocol.

Effects of foetal size, sex and developmental stage on adaptive transcriptional responses of skeletal muscle to intrauterine growth restriction in pigs.

Intrauterine growth restriction (IUGR) occurs both in humans and domestic species. It has a particularly high incidence in pigs, and is a leading cause of neonatal morbidity and mortality as well as impaired postnatal growth. A key feature of IUGR is impaired muscle development, resulting in decreased meat quality. Understanding the developmental origins of IUGR, particularly at the molecular level, is important for developing effective strategies to mitigate its economic impact on the pig industry and animal welfare. The aim of this study was to characterise transcriptional profiles in the muscle of growth restricted pig foetuses at different gestational days (GD; gestational length ~ 115 days), focusing on selected genes (related to development, tissue injury and metabolism) that were previously identified as dysregulated in muscle of GD90 fetuses. Muscle samples were collected from the lightest foetus (L) and the sex-matched foetus with weight closest to the litter average (AW) from each of 22 Landrace x Large White litters corresponding to GD45 (n = 6), GD60 (n = 8) or GD90 (n = 8), followed by analyses, using RT-PCR and protein immunohistochemistry, of selected gene targets. Expression of the developmental genes, MYOD, RET and ACTN3 were markedly lower, whereas MSTN expression was higher, in the muscle of L relative to AW littermates beginning on GD45. Levels of all tissue injury-associated transcripts analysed (F5, PLG, KNG1, SELL, CCL16) were increased in L muscle on GD60 and, most prominently, on GD90. Among genes involved in metabolic regulation, KLB was expressed at higher levels in L than AW littermates beginning on GD60, whereas both IGFBP1 and AHSG were higher in L littermates on GD90 but only in males. Furthermore, the expression of genes specifically involved in lipid, hexose sugar or iron metabolism increased or, in the case of UCP3, decreased in L littermates on GD60 (UCP3, APOB, ALDOB) or GD90 (PNPLA3, TF), albeit in the case of ALDOB this only involved females. In conclusion, marked dysregulation of genes with critical roles in development in L foetuses can be observed from GD45, whereas for a majority of transcripts associated with tissue injury and metabolism differences between L and AW foetuses were apparent by GD60 or only at GD90, thus identifying different developmental windows for different types of adaptive responses to IUGR in the muscle of porcine foetuses.

Involvement of miRNAs in equine follicle development.

Previous evidence from in vitro studies suggests specific roles for a subset of miRNAs, including miR-21, miR-23a, miR-145, miR-503, miR-224, miR-383, miR-378, miR-132, and miR-212, in regulating ovarian follicle development. The objective of this study was to determine changes in the levels of these miRNAs in relation to follicle selection, maturation, and ovulation in the monovular equine ovary. In Experiment 1, follicular fluid was aspirated during ovulatory cycles from the dominant (DO) and largest subordinate (S) follicles of an ovulatory wave and the dominant (DA) follicle of a mid-cycle anovulatory wave (n=6 mares). Follicular fluid levels of progesterone and estradiol were lower (P<0.01) in S follicles than in DO follicles, whereas mean levels of IGF1 were lower (P<0.01) in S and DA follicles than in DO follicles. Relative to DO and DA follicles, S follicles had higher (P≤0.01) follicular fluid levels of miR-145 and miR-378. In Experiment 2, follicular fluid and granulosa cells were aspirated from dominant follicles before (DO) and 24 h after (L) administration of an ovulatory dose of hCG (n=5 mares/group). Relative to DO follicles, L follicles had higher follicular fluid levels of progesterone (P=0.05) and lower granulosa cell levels of CYP19A1 and LHCGR (P<0.005). Levels of miR-21, miR-132, miR-212, and miR-224 were increased (P<0.05) in L follicles; this was associated with reduced expression of the putative miRNA targets, PTEN, RASA1, and SMAD4. These novel results may indicate a physiological involvement of miR-21, miR-145, miR-224, miR-378, miR-132, and miR-212 in the regulation of cell survival, steroidogenesis, and differentiation during follicle selection and ovulation in the monovular ovary.

Cardiac arrest following sugammadex administration.

A 68-year-old man underwent elective surgical repair of an abdominal wall hernia under general anaesthesia. The operation required muscle relaxation, for which we used rocuronium. Following completion of surgery, 180 mg sugammadex was administered intravenously. Shortly afterwards, the patient became severely bradycardic with hypotension, refractory to treatment with ephedrine. This progressed to a pulseless electrical activity cardiac arrest. After 4 min of cardiopulmonary resuscitation, there was return of spontaneous circulation and, following a period of haemodynamic stability in which general anaesthesia was maintained, the patient emerged from anaesthesia without incident. He remained haemodynamically stable until discharge. Post-resuscitation investigations including the serum tryptase level were unremarkable except for a mild respiratory acidosis and slightly elevated D-dimers. Sugammadex-induced bradycardia has previously been described, but its mechanism remains unknown. We believe that sugammadex was the cause of cardiac arrest in this case because of the timing and sequence of events, the evolution to pulseless electric activity and the relatively uneventful post-arrest clinical course. We hope that our report will help to promote awareness about this potential complication of a now commonly-used drug.

Contribution of local regeneration of glucocorticoids to tissue steroid pools.

11ß-Hydroxysteroid dehydrogenase 1 (11ßHSD1) is a drug target to attenuate adverse effects of chronic glucocorticoid excess. It catalyses intracellular regeneration of active glucocorticoids in tissues including brain, liver and adipose tissue (coupled to hexose-6-phosphate dehydrogenase, H6PDH). 11ßHSD1 activity in individual tissues is thought to contribute significantly to glucocorticoid levels at those sites, but its local contribution vs glucocorticoid delivery via the circulation is unknown. Here, we hypothesised that hepatic 11ßHSD1 would contribute significantly to the circulating pool. This was studied in mice with Cre-mediated disruption of Hsd11b1 in liver (Alac-Cre) vs adipose tissue (aP2-Cre) or whole-body disruption of H6pdh. Regeneration of [9,12,12-2H3]-cortisol (d3F) from [9,12,12-2H3]-cortisone (d3E), measuring 11ßHSD1 reductase activity was assessed at steady state following infusion of [9,11,12,12-2H4]-cortisol (d4F) in male mice. Concentrations of steroids in plasma and amounts in liver, adipose tissue and brain were measured using mass spectrometry interfaced with matrix-assisted laser desorption ionisation or liquid chromatography. Amounts of d3F were higher in liver, compared with brain and adipose tissue. Rates of appearance of d3F were ~6-fold slower in H6pdh-/- mice, showing the importance for whole-body 11ßHSD1 reductase activity. Disruption of liver 11ßHSD1 reduced the amounts of d3F in liver (by ~36%), without changes elsewhere. In contrast disruption of 11ßHSD1 in adipose tissue reduced rates of appearance of circulating d3F (by ~67%) and also reduced regenerated of d3F in liver and brain (both by ~30%). Thus, the contribution of hepatic 11ßHSD1 to circulating glucocorticoid levels and amounts in other tissues is less than that of adipose tissue.

Molecular characterization of five patients with homocystinuria due to severe methylenetetrahydrofolate reductase deficiency.

Methylenetetrahydrofolate reductase (MTHFR) plays a major role in folate metabolism. Disturbed function of the enzyme results in hyperhomocysteinemia and causes severe vascular and neurological disorders and developmental delay. Five patients suspected of having non-classical homocystinuria due to MTHFR deficiency were examined with respect to their symptoms, MTHFR enzyme activity and genotypes of the MTHFR gene. All patients presented symptoms of severe central nervous system disease. Two patients died, at the ages of 15 months and 14 years. One patient is currently 32 years old, and is being treated with betaine and folinic acid. The other two patients, with an early diagnosis and a severe course of the disease, are currently improving under treatment. MTHFR enzyme activity in the fibroblasts of four of the patients was practically undetectable. We found four novel mutations, three of which were missense changes c.664G> T (p.V218L), c.1316T> C (p.F435S) and c.1733T> G (p.V574G), and the fourth was the 1-bp deletion c.1780delC (p.L590CfsX72). We also found the previously reported nonsense mutation c.1420G> T (p.E470X). All the patients were homozygous. Molecular modelling of the double mutant allele (p.V218L; p.A222V) revealed that affinity for FAD was not affected in this mutant. For the p.E470X mutation, the evidence pointed to nonsense-mediated mRNA decay. In general, genotype-phenotype analysis predicts milder outcomes for patients with missense changes than for those in which mutations led to severe alterations of the MTHFR protein.

Relationships between size, steroidogenesis and miRNA expression of the bovine corpus luteum.

In a previous study, a subset of miRNAs were identified the expression of which increases substantially during the follicle-luteal transition in cattle. Here, we investigated the functional involvement of some of these miRNAs (miR-96, miR-182, miR-132, miR-21, miR-378) by determining whether there is an association in vivo between their expression in the corpus luteum (CL), CL size and progesterone production. The two largest and two smallest CL were collected from 12 donor beef heifers on Day 7 following ovarian super-stimulation (Day 0 = 28-32 h after first standing to be mounted). Additionally, the CL and a plasma sample were collected from 29 recipient heifers on Day 15. Luteal expression of miRNAs and mRNAs, and plasma progesterone concentrations were quantified by RT-qPCR and RIA, respectively. There were no differences in the mean expression of any miRNAs examined or the steroidogenic enzymes, STAR or CYP11A1, between the largest and smallest CL in donor heifers (P > 0.1). In addition, there were no significant correlations of luteal volume or weight with any miRNA, CYP11A1 or STAR in donor heifers. However, a correlation (r ≥ 0.5, P ≤ 0.001) existed between the transcript levels of CYP11A1 and STAR in the CL, as well as between each of those and miR-182 levels. In addition, CYP11A1 abundance was moderately correlated (r ≤ 0.4, P < 0.05) with each of miR-96 and miR-378. In recipient heifers, progesterone levels were moderately correlated with luteal weight (r = 0.41, P = 0.03) but not with the expression of any miRNA, CYP11A1 or STAR (P > 0.1). Moreover, luteal CYP11A1 and STAR were correlated (r = 0.6, P ≤ 0.001) with miR-182 as well as with each other, consistent with data in donor heifers. Finally, both CYP11A1 and STAR were moderately correlated (r ≤ 0.5) with miR-132 and, in the case of STAR, with miR-378. In summary, there was no association between either luteal weight/volume or plasma progesterone concentrations and any of the miRNAs analysed in donor and recipient heifers. However, CYP11A1 and STAR transcript levels were significantly correlated with several miRNAs, most notably miR-182, as well as with each other, in luteal tissues from both donor and recipient heifers. This finding confirms results of previous in vitro studies and, importantly, provides the first in vivo evidence of a role of the miR-183-96-182 cluster in regulating luteal steroidogenesis.

Calculation of MP2 and coupled-cluster molecular properties using the q-integral method.

The main purpose of this paper is to report results of quantum mechanical calculation of the H(2) system using the q-Integral method with correlation corrections to the SCF (Self Consistent Field) wave functions included through the Møller-Plesset second-order perturbation (MP(2)) and Coupled-Cluster (CC) theory. Using the q-Integral method, we evaluated potential energy curves, rovibrational spectroscopy constants, rovibrational spectra, interatomic equilibrium distance and longitudinal static hyper(polarizability). All calculations were carried out through the STO-3G, STO-6G, and double-zeta (DZV) atomic basis set. The q-Integral method was implemented in the source code of the general ab initio quantum chemistry package GAMESS.

Synthesis of d-glyco-alkynone derivatives via carbonylative Sonogashira reaction.

A carbonylative Sonogashira coupling approach to the synthesis of glyco-alkynones is described. Eighteen examples were obtained in moderate do nearly quantitative yields under mild conditions employing Mo(CO)6 as a safe carbon monoxide source. Functionalization of the alkynyl moiety via cycloaddition with organic azides provided six examples of glyco-triazoles.

Identification of Tobacco Types and Cigarette Brands Using an Electronic Nose Based on Conductive Polymer/Porphyrin Composite Sensors.

Three tobacco types (Burley, Flue Cured, and Oriental) and eight cigarette brands were unequivocally identified using an electronic nose formed by only three sensors based on a single novel conducting polymer (PF-BTB) doped with different porphyrins (H2TPP, H2TPFP, and H2BTBOP). The synthesis and characterization of the polymer are also discussed. Small changes in the porphyrin structure caused significant changes in the electrical conductance response patterns of the sensors upon exposure to complex chemical matrixes, representing a novel approach for tuning the selectivity of chemiresistive sensors for e-nose application. This e-nose is fast, cheap, reliable, can be easily operated, and could be a valuable tool for border agents fighting cigarette smuggling around the world, helping them prevent losses of millions in tax revenues and sales.

OBO-Protected Pyruvates as Reagents for the Synthesis of Functionalized Heteroaromatic Compounds.

Pd-catalyzed α-arylation of methyl-OBO-ketone (OBO = 4-methyl-2,6,7-trioxabicyclo[2.2.2]octan-1-yl) gives rise to arylated OBO-protected pyruvates. By appropriate prefunctionalization of the aryl ring or by subsequent functionalization at the α-carbonyl position of the arylated OBO-ketones, useful diketo OBO-protected carboxylates can be generated. Cyclization, aromatization, and OBO deprotection of these intermediates, using two distinct routes, gives access to valuable α-acyl heteroaromatic compounds.

Pyruvate Enolate Arylation and Alkylation: OBO Ester Protected Pyruvates as Useful Reagents in Organic Synthesis.

A protected pyruvate equivalent is described that allows arylation and arylation/alkylation reactions to be performed at the methyl group. Utilization of the OBO derivative of the pyruvate ester allowed the application of palladium catalyzed arylation reactions together with subsequent alkylation, under basic conditions. Moreover, the OBO protecting group could be easily removed in one step to provide access to a wide range of substituted pyruvate derivatives.

Effectiveness of percutaneous balloon mitral valvotomy during pregnancy.

During pregnancy, medically refractory congestive heart failure due to mitral stenosis continues to present a clinical challenge and optimal management remains controversial. Thirteen women underwent balloon mitral valvotomy for control of functional class III or IV congestive heart failure due to mitral stenosis during pregnancy. The mean gestational age at the time of valvotomy was 25 +/- 6 weeks. Percutaneous balloon mitral valvotomy was performed successfully in all patients. No maternal or fetal mortality occurred. The mean mitral valve area assessed by Doppler echocardiography increased from 0.9 +/- 0.3 cm2 before to 2.1 +/- 0.3 cm2 after valvotomy. The mean mitral valve gradient decreased from 20 +/- 7 to 4 +/- 2 mm Hg. This was associated with a decrease in the pulmonary artery systolic pressure from 62 +/- 24 to 32 +/- 14 mm Hg. Currently, 12 of the 13 patients have delivered at an average gestational age of 38 +/- 0.5 weeks. Symptoms of congestive heart failure improved in all women and all were in New York Heart Association functional class I at the time of delivery. One patient is still pregnant and symptom free. Eleven singlet pregnancies resulted in the birth of full-term, healthy infants (mean birth weight 3.2 kg). The woman carrying a twin pregnancy improved from New York Heart Association class IV to class I after balloon mitral valvotomy but delivered prematurely at 32 weeks. The premature twin infants weighed 1.0 and 1.5 kg and died from respiratory failure at 48 hours. Percutaneous balloon mitral valvotomy can be performed safely during pregnancy and is effective in relieving symptoms of severe congestive heart failure.(ABSTRACT TRUNCATED AT 250 WORDS)

B-CLL in PLL transformation associated with hypercalcemia.

A 64-year-old woman is reported with Stage I (Rai) chronic lymphocytic leukaemia (CLL), in whom hypercalcemia developed when an increased proportion of prolymphocytic cells characterized a transformation of CLL in prolymphocytoid leukaemia (CLL/PL). Although hypercalcemia is more frequently found in T-cell leukaemia associated with human T lymphotropic lymphocyte type I, scattered reports indicate that patients with B-CLL can also be affected with this metabolic disturbance. The case described here, progressed with an indolent course of CLL for 26 months, when she was admitted with a very aggressive disease characterized by a high WBC count, splenomegaly and hypercalcemia. Despite an effort to achieve a clinical remission, she failed treatment and death was attributed to unresponsive hypercalcemia. The mechanism of hypercalcemia in such cases is unclear as no parathyroid adenoma or second malignant tumor was found ante mortem. This electrolytic disturbance would appear to be a direct consequence of the transforming leukaemia and a possible mechanism involving a secreted humoral factor that could lead to altered calcium metabolism.

It is valid to dilate native aortic coarctation with a balloon catheter.

We report our experience in 37 patients with aortic coarctation who underwent balloon aortoplasty. Of the 37, the lesion was native in 34 cases. Clinical re-evaluation was possible in 22 patients, with a mean follow-up period of 13 months and, of these, 21 patients were asymptomatic. Haemodynamic and angiographic studies were performed in 13 patients, with a mean period of follow-up of 12 months, showing excellent results in 11 patients. Aneurysmal formation had occurred in one and recoarctation in the other patient. Our experience has proved that balloon aortoplasty is an effective method for treating patients with aortic coarctation. Adequate selection in terms of the morphology of the coarctation and the size of the balloon catheter are crucial factors in the success of the procedure.

Pulmonary valvoplasty--experience of 100 cases.

One-hundred patients with pulmonary valve stenosis underwent pulmonary valvoplasty, their ages ranging from 1 to 59 years. The systolic gradient across the valve ranged from 47 to 260 mm Hg (97.67 +/- 41.15) prior to the valvoplasty, and from 0 to 55 mm Hg (14.72 +/- 11.40) immediately after dilatation (P less than 0.0001). The clinical follow-up of 18.2 months of 56 patients showed a tendency for the systolic thrill to disappear, the systolic murmur became softer and there was a tendency to normalization of the electrocardiogram. A hemodynamic restudy was carried out in 54 patients and the systolic gradients across the valve ranged from 21.55 +/- 23.86. Seven patients required redilatation. In patients with hypertrophy of the infundibulum prior to valvoplasty this was observed to regress, indicating that infundibular hypertrophy is reversible.

[Pulmonary valvuloplasty with a balloon catheter in calcified pulmonary valve stenosis]. / Valvoplastia pulmonar com cateter balão na estenose pulmonar valvar calcificada.

Since the first pulmonary valvuloplasty report, the procedure has been used frequently, becoming the best option in isolated valvar pulmonary stenosis. Pulmonary valvuloplasty in adults with calcified valvar pulmonary stenosis, however, has been minimal. We have reported the case of a 62-year-old female patient, with calcified valvar pulmonary stenosis, who underwent valvuloplasty with balloon catheter and showed a decrease in the transvalvular systolic gradient, thus proving the success of post procedure.

[Results of percutaneous balloon valvuloplasty in membranous subaortic stenosis]. / Resultados da valvoplastia percutânea por cateter balão na estenose subaórtica em membrana.

PURPOSE: To evaluate the results of percutaneous balloon valvuloplasty (PBV) for membranous subaortic stenosis (MSAS). METHODS: Seven patients with MSAS were submitted to PBV using the retrograde approach. Hemodynamic and angiographic studies were performed before and after the procedure. The balloon/annulus ratio varied between 0.9 to 1.05. RESULTS: The systolic gradient (SG) between the left ventricle (LV) and aorta was reduced from 57.8 +/- 11.6 to 22.0 +/- 8.6 mmHg (p < 0.001) and the LV systolic pressure from 154.5 +/- 30.3 to 118.7 +/- 18.4 mmHg (p < 0.001). One patient had arterial occlusion and was submitted to surgical embolectomy and another one showed severe mitral regurgitation combined with aortic insufficiency (AI) and required surgical valvuloplasty. There was no mortality. Four patients had a 11 +/- 10 month follow up: the SG measured by echocardiogram was 34 +/- 7 mmHg and no patient showed worsening of the AI. CONCLUSION: PBV is an effective procedure to reduce the subaortic SG, with an acceptable morbidity and persistence of the results in the short term follow up. PBV is a valid alternative as an initial mode of therapy in the management of patients with MSAS.