Antonio Mucciolo

PPARγ in Placental Angiogenesis

Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor involved in diverse biological processes including adipocyte differentiation, glucose homeostasis, and inflammatory responses. Analyses of PPARγ knockout animals have been so far preempted by the early embryonic death of PPARγ-/- embryos as a consequence of the severe alteration of their placental vasculature. Using Sox2Cre/PPARγL2/L2 mice, we obtained fully viable PPARγ-null mice through specific and total epiblastic gene deletion, thereby demonstrating that the placental defect is the unique cause of PPARγ-/- embryonic lethality. The vasculature defects observed in PPARγ-/- placentas at embryonic d 9.5 correlated with an unsettled balance of pro- and antiangiogenic factors as demonstrated by increased levels of proliferin (Prl2c2, PLF) and decreased levels of proliferin-related protein (Prl7d1, PRP), respectively. To analyze the role of PPARγ in the later stage of placental development, when its expression peaks, we treated pregnant wild-type mice with the PPARγ agonist rosiglitazone. This treatment resulted in a disorganization of the placental layers and an altered placental microvasculature, accompanied by the decreased expression of proangiogenic genes such as Prl2c2, vascular endothelial growth factor, and Pecam1. Together our data demonstrate that PPARγ plays a pivotal role in controlling placental vascular proliferation and contributes to its termination in late pregnancy.

Transmission Fourier transform infrared microspectroscopy allows simultaneous assessment of cutin and cell-wall polysaccharides of Arabidopsis petals.

A procedure for the simultaneous analysis of cell-wall polysaccharides, amides and aliphatic polyesters by transmission Fourier transform infrared microspectroscopy (FTIR) has been established for Arabidopsis petals. The combination of FTIR imaging with spectra derivatization revealed that petals, in contrast to other organs, have a characteristic chemical zoning with high amount of aliphatic compounds and esters in the lamina and of polysaccharides in the stalk of the petal. The hinge region of petals was particular rich in amides as well as in vibrations potentially associated with hemicellulose. In addition, a number of other distribution patterns have been identified. Analyses of mutants in cutin deposition confirmed that vibrations of aliphatic compounds and esters present in the lamina were largely associated with the cuticular polyester. Calculation of spectrotypes, including the standard deviation of intensities, allowed detailed comparison of the spectral features of various mutants. The spectrotypes not only revealed differences in the amount of polyesters in cutin mutants, but also changes in other compound classes. For example, in addition to the expected strong deficiencies in polyester content, the long-chain acyl CoA synthase 2 mutant showed increased intensities of vibrations in a wavelength range that is typical for polysaccharides. Identical spectral features were observed in quasimodo2, a cell-wall mutant of Arabidopsis with a defect in pectin formation that exhibits increased cellulose synthase activity. FTIR thus proved to be a convenient method for the identification and characterization of mutants affected in the deposition of cutin in petals.

Is the GraftConnector a valid alternative to running suture in end-to-side coronary arteries anastomoses?

BACKGROUND An animal study was carried out to compare long-term patency rates of coronary anastomoses performed with the GraftConnector versus running suture technique. METHODS 10 sheep, 45 to 55 kg, underwent off-pump coronary artery bypass grafting (right internal mammary artery to left anterior descending artery). In 5 animals, the anastomosis was performed with a GraftConnector and in 5 animals with 7-0 running suture. Intraoperative fluoroscopy and a fluoroscopic control at 6 months were performed. After 6 months, the animals were sacrificed and the anastomoses were examined histologically. RESULTS All animals survived at 6 months with 100% anastomosis patency rates in both groups. In the GraftConnector group, the anastomosis diameter at 6 months fluoroscopy was 118% of native left anterior descending artery versus 97% of the control group. Luminal anastomotic width at histology was 1.7 +/- 0.2 mm in the device group versus 1.6 +/- 0.1 mm in the control group. Mean intimal hyperplasia thickness was 0.21 +/- 0.1 mm in the device group versus 0.01 mm in the control group. CONCLUSIONS The GraftConnector provides a consistent and reproducible coronary artery anastomosis and reduces technical demand and manual dexterity in coronary operations. Long-term results demonstrate that off-pump coronary artery bypass grafting performed with the GraftConnector had the same patency rate and luminal width as those performed with running suture.

The ABCG transporter PEC1/ABCG32 is required for the formation of the developing leaf cuticle in Arabidopsis

The cuticle is an essential diffusion barrier on aerial surfaces of land plants whose structural component is the polyester cutin. The PERMEABLE CUTICLE1/ABCG32 (PEC1) transporter is involved in plant cuticle formation in Arabidopsis. The gpat6 pec1 and gpat4 gapt8 pec1 double and triple mutants are characterized. Their PEC1-specific contributions to aliphatic cutin composition and cuticle formation during plant development are revealed by gas chromatography/mass spectrometry and Fourier-transform infrared spectroscopy. The composition of cutin changes during rosette leaf expansion in Arabidopsis. C16:0 monomers are in higher abundance in expanding than in fully expanded leaves. The atypical cutin monomer C18:2 dicarboxylic acid is more prominent in fully expanded leaves. Findings point to differences in the regulation of several pathways of cutin precursor synthesis. PEC1 plays an essential role during expansion of the rosette leaf cuticle. The reduction of C16 monomers in the pec1 mutant during leaf expansion is unlikely to cause permeability of the leaf cuticle because the gpat6 mutant with even fewer C16:0 monomers forms a functional rosette leaf cuticle at all stages of development. PEC1/ABCG32 transport activity affects cutin composition and cuticle structure in a specific and non-redundant fashion.

Biologic Fixation of Polyester-versus Polyurethane-covered Stents in a Porcine Model

PURPOSE Migration of endoprostheses remains a concern in endovascular aneurysm treatment. Biologic fixation is supposed to enhance anchorage, but the diseased atherosclerotic aorta in humans has demonstrated a limited capacity to incorporate an endoprosthesis by cellular proliferation. The biologic response of two different types of endoprostheses was evaluated in the porcine aorta. MATERIALS AND METHODS Two types of endoprostheses--four polyurethane-covered (PUC) stents with a macroporous polyurethane covering and four polyester-covered (PEC) stents with a woven polyester covering--were implanted in eight infrarenal porcine aortas for 6 weeks. Electron microscopy and qualitative and quantitative microscopy were performed on serial cross sections. RESULTS The PUC stents demonstrated an increase in diameter (from 8 mm +/- 1 to 10 mm +/- 1, 12.5%; P = .009), whereas the PEC stents persisted in their original dimensions (8 mm +/- 1, 0%). PUC and PEC stents were covered by continuous thrombus-free neointima (269 microm +/- 51 vs 575 microm +/- 113, respectively; P < .01). The PUC stents demonstrated firm attachment to the aorta as a consequence of a granulation tissue with ingrowth into the pores of the polyurethane covering. The PEC stents remained in loose contact with the aorta in the absence of tissue ingrowth. CONCLUSIONS Enhanced biologic fixation was achieved by extensive granulation tissue invading the pores of PUC endoprostheses. This finding can modify the design of future devices.

A prototype paediatric venous cannula with shape change in situ

During cardiopulmonary bypass (CPB), venous drainage may be impeded due to small vessel and cannula size or chattering, thus, blood return to the heart-lung machine is reduced. We designed a self-expandable prototype cannula, which is able to maintain the vein open and overcome this problem and analysed its performance capability. This prototype and several other cannulae were tested using an access vessel diameter of 7 mm. An in vitro circuit was set up with a 10 mm penrose latex tube simulating the patient’s vein placed between the patient preload reservoir and the cannula, encasing the cannula’s inlet(s). Maximum flow rate was determined for passive venous drainage (PVD) at preloads (P) of 2 and 4 mmHg. We compared these results to three classic single-stage venous cannulae: basket tip, thoracic drain and percutaneous tip. By comparing the other cannulae to the prototype, under PVD conditions and a central venous pressure (CVP) of 2 mmHg, the prototype cannula’s flow rate (1.329 /0.04 L/min) outperformed the basket type (the best performing comparator) (1.029 / 0.08 L/min) by 23% (p B /0.005). When the preload was increased to 4 mmHg under PVD conditions, the same trend was noted with the prototype cannula (1.659 /0.05 L/min), outperforming the basket cannula’s value (1.269 /0.05 L/min) by 24% (p B /0.001). This new cannula design provides superior flow characteristics, under all test conditions, compared to the classic single-stage venous cannulae used for paediatric CPB surgery.

Connecting the Molecular Structure of Cutin to Ultrastructure and Physical Properties of the Cuticle in Petals of Arabidopsis

GPAT6 and DCR play different roles in structuring the cell wall-cuticle continuum. The plant cuticle is laid down at the cell wall surface of epidermal cells in a wide variety of structures, but the functional significance of this architectural diversity is not yet understood. Here, the structure-function relationship of the petal cuticle of Arabidopsis (Arabidopsis thaliana) was investigated. Applying Fourier transform infrared microspectroscopy, the cutin mutants long-chain acyl-coenzyme A synthetase2 (lacs2), permeable cuticle1 (pec1), cyp77a6, glycerol-3-phosphate acyltransferase6 (gpat6), and defective in cuticular ridges (dcr) were grouped in three separate classes based on quantitative differences in the ν(C=O) and ν(C-H) band vibrations. These were associated mainly with the quantity of 10,16-dihydroxy hexadecanoic acid, a monomer of the cuticle polyester, cutin. These spectral features were linked to three different types of cuticle organization: a normal cuticle with nanoridges (lacs2 and pec1 mutants); a broad translucent cuticle (cyp77a6 and dcr mutants); and an electron-opaque multilayered cuticle (gpat6 mutant). The latter two types did not have typical nanoridges. Transmission electron microscopy revealed considerable variations in cuticle thickness in the dcr mutant. Different double mutant combinations showed that a low amount of C16 monomers in cutin leads to the appearance of an electron-translucent layer adjacent to the cuticle proper, which is independent of DCR action. We concluded that DCR is not only essential for incorporating 10,16-dihydroxy C16:0 into cutin but also plays a crucial role in the organization of the cuticle, independent of cutin composition. Further characterization of the mutant petals suggested that nanoridge formation and conical cell shape may contribute to the reduction of physical adhesion forces between petals and other floral organs during floral development.

Cuticular Defects in Oryza sativa ATP-binding Cassette Transporter G31 Mutant Plants Cause Dwarfism, Elevated Defense Responses and Pathogen Resistance

The cuticle covers the surface of the polysaccharide cell wall of leaf epidermal cells and forms an essential diffusion barrier between plant and environment. Homologs of the ATP-binding cassette (ABC) transporter AtABCG32/HvABCG31 clade are necessary for the formation of a functional cuticle in both monocots and dicots. Here we characterize the osabcg31 knockout mutant and hairpin RNA interference (RNAi)-down-regulated OsABCG31 plant lines having reduced plant growth and a permeable cuticle. The reduced content of cutin in leaves and structural alterations in the cuticle and at the cuticle-cell wall interface in plants compromised in OsABCG31 expression explain the cuticle permeability. Effects of modifications of the cuticle on plant-microbe interactions were evaluated. The cuticular alterations in OsABCG31-compromised plants did not cause deficiencies in germination of the spores or the formation of appressoria of Magnaporthe oryzae on the leaf surface, but a strong reduction of infection structures inside the plant. Genes involved in pathogen resistance were constitutively up-regulated in OsABCG31-compromised plants, thus being a possible cause of the resistance to M. oryzae and the dwarf growth phenotype. The findings show that in rice an abnormal cuticle formation may affect the signaling of plant growth and defense.

Does large oversizing of self-expandable endoprostheses compensate for aortic growth?

PURPOSE There is experimental evidence that self-expandable endoprostheses are appropriate for compensation of aortic growth. A potential future application includes their use in the treatment of aortic coarctation. Yet their behavior is poorly investigated. The present study evaluates the performance of largely oversized self-expandable endoprostheses (EPs) in the growing porcine aorta and the biologic response toward them. METHODS Twenty oversized EPs (Talent, TalentLoPro, Stenway, and Wallstent; nominal lumen area 314 mm(2) [diameter 20 mm]) were implanted in the descending thoracic aorta of juvenile pigs. Four nonoversized EPs (TalentLoPro; nominal lumen area 154 mm(2) [diameter 14 mm]) served as controls. Cross sections of aorta and EPs were measured by intravascular ultrasound (IVUS) at implantation and 3 months follow-up. The histologic response was assessed by microscopy. RESULTS At implantation, the EPs were 99 +/- 41% oversized relative to the aortic area [48 +/- 22% in diameter]. At follow-up, the area of the aortic lumen increased 60 +/- 50%, P <.001 [29 +/- 23% in diameter]. The Stenway EPs demonstrated the largest prosthetic lumen with 80 +/- 10% of their nominal area (P =.001). However, the prosthetic lumen area was reduced by intimal hyperplasia in all devices. The largest perfused lumen was obtained by the TalentLoPro EPs due to their progressive expansion. Nonoversized EPs resulted in a significantly smaller perfused lumen (P =.012). The inflammatory response to polyester was moderate, whereas polyurethane evoked an extensive body-foreign reaction. CONCLUSION Large oversizing of self-expandable EPs is essential in order to maintain a large perfused lumen during aortic growth, thereby minimizing the adverse effect of intimal hyperplasia. This was best achieved by the TalentLoPro EPs.

The GraftConnector experience. Long-term patency and histological work up in an animal model.

BACKGROUND A device to perform sutureless end-to-side coronary artery anastomosis has been developed by means of stent technology (GraftConnector). The present study assesses the long-term quality of the GraftConnector anastomosis in a sheep model. METHODS In 8 adult sheep, 40-55 kg in weight, through left anterior thoracotomy, the right internal mammary artery (RIMA) was prepared and connected to the left anterior descending artery (LAD) by means of GraftConnector, on beating heart, without using any stabilizer. Ticlopidine 250 mg/day for anticoagulation for 4 weeks and Aspirin 100 mg/day for 6 months were given. The animals were sacrificed after 6 months and histological examination of anastomoses was carried out after slicing with the connector in situ for morphological analysis. RESULTS All animals survived at 6 months. All anastomoses were patent and mean luminal width at histology was 1.8 +/- 0.2 mm; mean myotomia hyperplasia thickness was 0.21 +/- 0.1 mm. CONCLUSIONS Long-term results demonstrate that OPCABGs performed with GraftConnector had 100% patency rate. The mean anastomotic luminal width corresponds to mean LADs adult sheep diameter. We may speculate that myotomia hyperplasia occurred as a result of local device oversizing.