Plant-Herbivore and Plant-Pollinator Interactions of the Developing Perennial Oilseed Crop, Silphium integrifolium.

Sampling in Kansas and North Dakota documented the plant-herbivore and plant-pollinator interactions of the developing perennial oilseed crop, Silphium integrifolium Michx. The larva of the tortricid moth, Eucosma giganteana (Riley), was the most damaging floret- and seed-feeding pest in Kansas, with infested heads producing ≈85% (2015) or ≈45% (2016) fewer seeds than apparently undamaged heads. Necrosis of apical meristems caused stunting and delayed bloom in Kansas; though the source of the necrosis is not known, observations of the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois; Hemiptera: Miridae), in S. integrifolium terminals suggest a possible cause. In North Dakota, E. giganteana larvae were not found, but pupae of Neotephritis finalis (Loew; Diptera: Tephritidae), a minor pest of cultivated sunflower, were common in the heads of S. integrifolium. Bees appeared highly attracted to S. integrifolium, and in all but one observation, bees were seen actively collecting pollen. The most common bees included large apids (Apis mellifera L., Svastra obliqua [Say], Melissodes spp.) and small-bodied halictids (Lasioglossum [Dialictus] spp.). Controlled pollination experiments demonstrated that S. integrifolium is pollinator dependent, due to both mechanical barriers (imperfect florets and protogyny) and genetic self-incompatibility. Subsequent greenhouse tests and AFLP confirmation of putative self-progeny show that a low (<1%) level of self-pollination is possible. If genetic self-incompatibility is eventually reduced through breeding, mechanical barriers would maintain a reliance on bees to move pollen between male and female florets. Collectively, observations on S. integrifolium show that both herbivore and pollinator management are important to maximize seed production.

Small plot trials documenting effective mating disruption of oriental fruit moth by using high densities of wax-drop pheromone dispensers.

In 2004 field experiments, we compared the effectiveness of various deployment densities of 0.1-ml paraffin wax drops containing 5% pheromone versus Isomate M-Rosso "rope" dispensers for disruption of Grapholita molesta (Busck). Treatments were evaluated in 0.05-ha (12-tree) plots of 'Delicious' apples receiving regular maintenance according to growers' standards, but not sprayed with insecticides. The application densities of 0.1-ml wax drops were 3 per tree (820/ha), 10 per tree (2,700/ha), 30 per tree (8,200/ha), and 100 per tree (27,300/ha). Wax drops were compared with 3-ml dispensers of pheromone-containing paraffin wax or Isomate M-Rosso ropes at 1.8 per tree (500/ha) and untreated control plots. Treatments were applied before the start of each of three moth generations. Orientational disruption, as measured by inhibition of moth captures in pheromone-baited delta traps, was greatest in plots that received 100 drops per tree (99.2%) and 30 drops per tree (99.4%). More than 55% of tethered, virgin females were mated in control plots after one night of deployment. However, no mating was recorded at the two highest application densities of wax drops where orientational disruption of traps exceeded 99%. Mating ranged from 7 to 20% among the other treatments, including Isomate rope dispensers. G. molesta males were observed closely approaching pheromone dispensers in plots containing ropes and wax drops, documenting competitive attraction between synthetic pheromone sources and feral females. The majority of observed G. molesta males approached within 60 cm of wax drops or pheromone ropes and departed within 20 s by flying upwind. Thirty wax drops per tree yielded higher mating disruption of G. molesta than did Isomate M-Rosso dispensers deployed at the recommended rate of 500/ha (1.8 per tree). Measurement of release rates confirmed behavioral data indicating that paraffin wax dispensers would need to be applied once per G. molesta generation in Michigan. Paraffin wax drops are a promising technology for moth mating disruption. They are cheaper and easier to produce, require less total pheromone per annual application, and produce better mating disruption at appropriate deployment densities compared with Isomate M-Rosso dispensers under high G. molesta population densities. The cost-effectiveness of this approach will require an appropriate mechanized applicator for wax drops.

Diltiazem during reperfusion preserves high energy phosphates by protection of mitochondrial integrity.

OBJECTIVE: This study evaluates the effects of diltiazem administered during reperfusion on hemodynamic, metabolic, and ultrastructural postischemic outcome. METHODS: Hearts of 38 adult White New Zealand rabbits underwent 60 min of global cold ischemia followed by 40 min of reperfusion in an erythrocyte perfused isolated working heart model. Hearts were randomly assigned to four groups and received diltiazem (0.1, 0.25, and 0.5 micromol/l) during reperfusion only, or served as control. RESULTS: The postischemic time courses of heart rate, aortic flow, and external stroke work clearly reflected the dose-dependent negative chronotropic and inotropic efficacy of diltiazem in the two higher concentrations. High energy phosphates (HEP) determined from myocardial biopsies taken after 40 min of reperfusion were significantly better preserved in all treatment groups compared to control hearts. Similarly ultrastructural grading of mitochondria and myofilaments revealed a significant reduction of reperfusion injury in hearts that received diltiazem compared to control. CONCLUSIONS: Diltiazem protects mitochondrial integrity and function, thereby preserving myocardial HEP levels. Only low dose diltiazem (0.1 micromol/l) during reperfusion combines both, optimal mitochondrial preservation with minimal changes in hemodynamics.

The differentiation inducers phenylacetate and phenylbutyrate modulate camptothecin sensitivity in colon carcinoma cells in vitro by intracellular acidification.

Aromatic fatty acids such as phenylbutyrate (PB) and its metabolite phenylacetate (PA) induce growth arrest, differentiation and apoptosis in solid tumor cells. Despite their antiproliferative action they were reported to exhibit a synergistic effect in combination with cytotoxic drugs like topotecan, and others. Since the activity of the camptothecines (CPTs) depends on local pH conditions, we investigated, whether PB/PA modulate CPT effects indirectly by affecting intracellular pH in SW620 and SW480 colon cancer cells. The results for the colon carcinoma cells show an antagonistic interaction for the combination of CPT and 0.25-5 mM PA in viability assays, resulting in an approximately 3-fold increase in IC50 (control: 20+/-7 nM). A synergistic effect with significantly increased numbers of late apoptotic/necrotic cancer cells (difference +21+/-4%) and 1.4-fold sensitization were detected upon inclusion of 2.5 mM PA during a 4-h CPT (10 micro;M) loading phase. In response to 0.25-1 mM PA/PB the cells exhibit a reversible decrease of pHi (0.1-0.31 pH units) in HEPES- or bicarbonate-buffered media. Dose-dependent acidification and pHi-recovery occurred following addition of PA and PB after an acid load and inhibition of the Na+/H+-antiporter and bicarbonate exchangers, pointing to a possible intracellular mechanism of cytoplasmic acidification. It is concluded that the synergistic modulation of CPT toxicity by short-term PA/PB treatment in colon carcinoma cells is caused by changes in intracellular pH, possibly affecting quantity and localization of the active closed lactone form of this drug.

Light and electron microscopic study of stress-shielding effects on rat patellar tendon.

In this second part of our study, the histomorphologic changes occurring in the patellar tendon (PT) of rats after sole stress-shielding were evaluated. In seven adult albino rats, both PTs were exposed by straight skin incision and then stress-shielded on one side by a cerclage, while the contralateral PT served as the sham-operated control. One animal died after the operation and was used as a negative control. After 10 weeks of otherwise unrestricted motion, the rats were killed, and the histomorphology of all PT specimen pairs compared by light and transmission electron microscopy. Light microscopy showed mid-portion thickening and irregularity of collagen bundles in the stress-shielded tendons. Intense remodelling was demonstrated by increased cellularity and vascularity, as well as by enrichment in acidic proteoglycans. Ultrastructural evaluation and morphometry revealed a predominance of large diameter (peak between 180 and 260 nm) collagen fibrils in the sham-operated controls, while in the stress-shielded tendons the number of apparently new, small-diameter (peak between 40 and 60 nm) collagen fibrils increased (up to 77% per cross-sectional field of view). The difference in peak diameters was statistically significant (p < 0.0005). This rat model demonstrated that sole stress-shielding not only causes biomechanical alterations, but also intense tissue remodelling and significant morphological changes in the collagen fibrils in the patellar tendon, comparable to so-called 'ligamentization' in experimental and clinical patellar tendon grafts for anterior cruciate ligament reconstruction.

Modulation of CD99/MIC2 expression of human AHTO-7 osteoblasts by carcinoma cell line-conditioned media.

BACKGROUND: Prostate cancer metastases induce a predominantly osteoblastic response in bone tissue, resulting in new bone formation and associated morbidity; however, the mechanisms of these tumor-host responses are not fully understood. MATERIALS AND METHODS: Supernatants of prostate (PC3, DU145, LNCaP), breast (BT20, ZR-75-1), colon (SW620, Colo 320DM), pancreatic (ASPC1, Capan-1), renal cell (ACHN) and hepatoma (HepG2) cell lines were tested for their capacity to modulate proliferation, activity of alkaline phosphatase (ALP) and CD99/MIC2 expression in AHTO-7 (large T antigen transfected human trabecular osteoblasts) cells in vitro. RESULTS: Osteoblastic stimulation is not restricted to prostate cancer derived conditioned media CM and high activity is found in CM from Capan-1, HepG2 and ACHN lines. Furthermore these CM down-regulate the expression of the CD99/MIC2 antigen in comparison to medium by AHTO-7 cells as detected by HBA-71 immunofluorescence, with the exception of prostate cancer-derived CM. Induction of the differentiation marker ALP was detected in response to CM derived from Capan-1, BT-20 and Colo320DM. Stimulation of the proliferation of AHTO-7 cells (105-138% of control), induction of ALP (1.17-5.29-fold) and down-regulation of CD99 (13.6-57.5%) exhibited no correlation. CM derived from PC3 and LNCaP metastatic prostate cancer cell lines specifically resulted in the retention/stimulation of the expression of CD99/MIC2 in AHTO-7 cells in contrast to all other cell lines tested. CONCLUSION: The CD99/MIC2 antigen, which is expressed on human osteoblasts and osteosarcoma cells, seems to constitute a new and independent response marker of osteoblasts in the triggering of osteoblastic reaction by prostate cancer cells.

Pre- and early postoperative risk factors for death after cardiac transplantation: a single center analysis.

Due to the limited number of donor organs, death on the waiting list and waiting time for cardiac transplantation have markedly increased. A pressing need of appropriate selection criteria for patients who would benefit most from transplantation is apparent. The purpose of this study is to identify pre- and early postoperative risk factors that influence long term survival after cardiac transplantation. 702 consecutive patients who underwent cardiac transplantation between 3/1984 and 12/1997 were analyzed retrospectively for the influence of different pre- and early postoperative risk factors on early (30 days) and late death (5 years). Univariate and multivariate regression analysis revealed risk factors for early as well as late death. Predictors of early death were higher preoperative PVR, retransplantation, longer ischemic time, postoperative acute kidney failure and longer intubation time. Risk factors for late death were early transplant era, previous cardiac surgery, patients awaiting transplantation in a hospital, prolonged stay in an intensive care unit, and any rejection during the first month after transplantation. These results demonstrate that pre- and early postoperative risk factors have significant influence on early and long term survival.

Morphologic changes correlating to different sensitivities of Escherichia coli and enterococcus faecalis to Nd:YAG laser irradiation through dentin.

BACKGROUND AND OBJECTIVE: Previous studies demonstrated the disinfecting potential of Nd:YAG laser irradiation on the root canal system from an overall quantitative viewpoint. The aim of this study was to evaluate the specific effect of irradiation through dentin on gram-negative and gram-positive bacteria with regard to their cell structure. STUDY DESIGN/MATERIALS AND METHODS: Sterile dentin samples of standardized size were divided into two sets of four groups with eight samples each. The first set was inoculated with Escherichia coli as the gram-negative test strain, the second set was inoculated with Enterococcus faecalis, which served as the gram-positive test organism. The samples were then irradiated on the bacteria-free side in contact mode under constant scanning movement at an angle of 10 degrees by use of the fiber optic of the Nd:YAG laser. Upon laser treatment they were critical point dried and subjected to SEM investigation. Another two sets of samples were prepared and irradiated in the same manner and evaluated by standard microbiological procedures to verify whether the observed morphologic alterations correlated to cell death. RESULTS: SEM investigations revealed damage pattens that increased with the amount of energy applied. Whereas the gram-negative test organism showed immediate structural injury, the gram-positive test organism required repeated application of irradiation. The microbiological examination showed reduction of both bacterial strains, yet to different extents. CONCLUSION: Our study demonstrates the different morphologic impact of Nd:YAG laser irradiation through dentin on representatives of the two main groups of bacteria. It shows that the construction of the cell wall is crucial for their individual sensitivity to laser treatment.

Resveratrol pretreatment desensitizes AHTO-7 human osteoblasts to growth stimulation in response to carcinoma cell supernatants.

Resveratrol, a natural phytoestrogen, has been reported to promote differentiation of murine MC3T3-E1 osteoblasts and to inhibit proliferation of prostate cancer cell lines. In the present study we tested the effects of resveratrol on the increased proliferation of human AHTO-7 osteoblastic cell line induced by conditioned media (CM) from a panel of carcinoma cell lines. This compound was found to modulate AHTO-7 proliferation in a tamoxifen-sensitive mechanism at lower concentrations, but failed to induce the osteoblast differentiation marker alkaline phosphatase (ALP) in contrast to vitamin D3. The proliferative response of AHTO-7 cells to conditioned media from carcinoma cell lines was diminished (30-71.4% inhibition) upon pretreatment with 0.5 microM resveratrol. Highest inhibition was demonstrated for pancreas (BxPC3, Panc-1), breast (ZR75-1) and renal (ACHN) carcinoma cell line supernatants whereas the effect on colon carcinoma (SW620, Colo320DM) cell CM and prostate cancer (PC3, DU145 and LNCaP) CM was less pronounced. Direct addition of resveratrol affected only supernatants of cell lines (<25% inhibition) exhibiting growth stimulatory activity for normal WI-38 lung fibroblasts. Resveratrol inhibited proliferation of DU145 and LNCaP cells in concentrations exceeding 5 microM, altered cell cycle distribution of all prostate cancer cell lines in concentrations as low as 0.5 microM, but did not inhibit the production of osteoblastic factors by these lines. In conclusion, resveratrol failed to induce ALP activity as marker of osteoblast differentiation in human osteoblastic AHTO-7 cells, however, inhibited their response to osteoblastic carcinoma-derived growth factors in concentrations significantly lower than those to reduce growth of cancer cells, thus effectively modulating tumor - osteoblast interaction.

Effect of trephination technique on the ultrastructure of corneal transplants: guided trephine system v posterior punch technique.

AIM: Different trephination methods may lead to differences in degree of tissue damage and endothelial cell loss, which both influence the outcome of penetrating keratoplasty. Light, transmission, and scanning electron microscopy were used to compare the ultrastructural appearance of the cut edges and the endothelial cell loss in 26 human corneal donor buttons obtained by trephination with the suction fixated guided trephine system (GTS) and with the free hand posterior punch technique (PPT). METHODS: Human corneas were stored between 5 and 14 days in Optisol. One cornea from each pair was used for each technique. Trephinations (7.5 mm) were performed either from the anterior direction with the GTS (n=13) or from the posterior direction with the PPT (n=13) using Pharmacia Superblade trephines. Light microscopy, transmission electron, and scanning electron microscopy were performed according to standard procedures. Widening of the cut edges and the extent of endothelial cell loss were measured at three different areas per corneal button and analysed statistically. RESULTS: In contrast with the PPT, the GTS trephine produced considerable fibrillar disorder at the cut edges of the corneal buttons. The distance to which the endothelial cell loss extended from the edges of the cuts was significantly (p<0. 001) lower for the GTS (42.2 (SD 50.8) microm from the edge) than for the PPT (109.3 (68.1) microm). Stromal widening at the edges (measured as percentage increase in stromal thickness, compared with the thickness of the central cornea) was observed with both techniques. However, the mean stromal widening produced by the GTS was significantly greater than that produced by PPT (106% (24%) v 69% (21%); p<0.002). CONCLUSION: Both trephination techniques produced only minor tissue damage. Nevertheless, there were distinct differences in the fine appearance of the cuts produced by the GTS and the PPT techniques. The extent of the fibrillar dislocation and stromal widening was greater at the edges of the GTS buttons. The GTS technique produced significantly less endothelial cell loss at the cut edges than did the free hand punching technique, PPT.

A simple HPLC method for monitoring mycophenolic acid and its glucuronidated metabolite in transplant recipients.

Mycophenolic acid (MPA) is nowadays in broad clinical use as a substitute for azathioprine. An immunoassay for MPA recently received approval for clinical applications. The high performance liquid chromatography (HPLC) assay for measuring MPA and its glucuronide conjugate (MPAG) we describe here is not only rapid and simple but also extremely sensitive at plasma levels obtained during standard immunosuppressive regimens. The determination of MPAG is possible without any change of the chromatographic conditions (detection wavelength of 214 nm, mobile phase: acetonitrile and 50 mmol/l o-phosphoric acid (50:50, V/V), run time: 15 min). The required equipment is a standard HPLC system including a simple UV-detector. Sample volume of 400 microl is required for both determinations. Detection limit is 0.25 micromol/l for MPA and 5 micromol/l for MPAG. Linearity is excellent for serial dilutions (0.5-25 micromol/l for MPA, 25-500 micromol/l for MPAG) and high accuracies favour the method described. More than 2000 plasma samples tested for MPA in patients after heart transplantation within one year and more than 500 samples for MPAG underline the clinical applicability of this assay.

Ketoprofen-poly(D,L-lactic-co-glycolic acid) microspheres: influence of manufacturing parameters and type of polymer on the release characteristics.

The effect of manufacturing parameters on the size and drug-loading of ketoprofen-containing biodegradable and biocompatible poly(DL-lactic-co-glycolic acid) (PLGA) microspheres prepared by the solvent evaporation method was investigated. For both drug-free and drug-loaded microspheres, smaller microspheres with a narrower size distribution were obtained when the stirring rate or the volume of the organic phase was increased. Incorporation of ketoprofen was found to increase with increasing volume of the organic phase and decreasing pH of the aqueous phase, but was independent of the acidity and the inherent viscosity of the PLGA used. The biphasic release profile of ketoprofen from the microspheres was dependent on the type of PLGA as well as the size and drug-loading, two parameters governed by the manufacturing process. The first burst effect was found to increase with the drug content, reduction of size of the microspheres and increasing inherent viscosity of the matrix, whereas acidity of the PLGA had no effect on the release of this acidic drug. A vigorous first burst effect was associated with reduced sustained delivery of ketoprofen, the rate of the delayed release phase being dependent on the inherent viscosity of the matrix, the size, the payload and the pH during preparation of the microspheres. Thus, by selection of the manufacturing parameters and the type of PLGA, it is possible to design a controlled drug delivery system for the prolonged release of ketoprofen, improving therapy by possible reduction of time intervals between peroral administration and reduction of local gastrointestinal side effects.

Interlacing and cross-angle distribution of collagen lamellae in the human cornea.

PURPOSE: The interlacing and cross angles between the collagen lamellae within the human corneal stroma were studied by means of scanning electron microscopy (SEM). METHODS: For SEM, cells and noncollagenous extracellular matrix were removed with 10% sodium hydroxide. Transmission electron microscopy (TEM) preparations were performed according to standard procedures. The interlacing of lamellae was studied within the limbal, paracentral, and central regions of five different layers. The cross angles between the longitudinal axes of adjacent lamellae were measured. The distribution of these angles within defined layers and regions was compared. Special attention was paid to the interlacing of the lamellae. RESULTS: Lamellae split in an anteroposterior direction as well as horizontally into branches and are interlaced by crossing the fissures between the branches. Smaller lamellae cross through clefts of neighboring lamellae. The cross angles show a high variability of 1 degree - 90 degrees. With the exception of the limbal region of the layer adjacent to Descemet's membrane, the distribution of cross angles is similar. A frequent occurrence of cross angles <30 degrees (68%) in this limbal layer can be explained by a pseudocircular orientation (ligamentum circulare corneae) of the lamellae. CONCLUSION: The present study shows that the three-dimensional organization of the collagen lamellae is characterized by a greater extent of lamellar interlacing than has been assumed until now.

Altered organization of collagen in the apex of keratoconus corneas.

PURPOSE: In 15 keratoconus corneas, the three-dimensional arrangement of collagen lamellae was investigated by means of scanning electron microscopy. METHODS: Keratoconus corneas without visible scars were obtained during perforating keratoplasty. The noncollagenous matrix of the stroma was removed with sodium hydroxide. Descemet's membrane was removed mechanically and deeper layers of the stroma were exposed by cutting the tissue tangentially to the corneal surface with an ultramicrotome. The apical and the para-apical regions of keratoconus were compared the central regions of normal corneas. RESULTS: In the apical regions of 11 out of the 15 keratoconus corneas (73%), the arrangement of the collagen lamellae differs from those of the para-apical regions and normal corneas. Their collagen fibrils from uniform layers and no delimited collagen lamellae can be differentiated. Interlacing between adjacent layers in extremely decreased or even absent. In the para-apical region of keratoconus corneas the three-dimensional arrangement of collagen lamellae does not differ from that in normal corneas. CONCLUSION: Stromal thinning and conical ectasia in the apex of keratoconus corneas alters the organization of collagen. This will certainly affect the biomechanical properties of the cornea and further lead to a progression of keratoconus irrespective of its primary pathogenesis.

Ultrastructure of clear corneal incisions. Part II: Corneal trauma after lens implantation with the Microstaar injector system.

PURPOSE: To examine the ultrastructure of clear corneal incisions (CCIs) after implantation of a plate-hepatic intraocular lens (IOL) with the Microstaar injector system through two incision widths. SETTING: University Eye Clinic and Institute of Histology and Embryology II, University of Vienna, Austria. METHODS: Fourteen human cadaver eyes without prior ocular surgery were obtained from the University Eye Bank, Vienna. Single-plane CCIs were performed with 3.0 and 3.2 mm steel keratomes. Using the Microstaar injector system, a foldable silicone plate-haptic IOL (23 diopters) was implanted in the anterior chamber. During the entire procedure, the eye pressure was kept between 26 and 30 mm Hg by infusing balanced salt solution into the anterior chamber. Specimens for light microscopy and scanning electron microscopy were prepared according to standard procedures. RESULTS: After IOL implantation, the 3.0 mm steel blade incisions exhibited distinct distensions at their lateral ends. Adjacent to these distensions, the collagen lamellae were pressed apart, displaced, and torn. In 3.2 mm tunnels, the corneal trauma at both lateral ends was considerably less severe. These incisions also showed a better primary adaptation of the wound lips after implantations. CONCLUSIONS: Implantation performed with the Microstaar injector system through 3.0 mm steel blade CCIs led to considerably more severe corneal trauma than implantation through 3.2 mm incisions.

Ultrastructure of clear corneal incisions. Part I: Effect of keratomes and incision width on corneal trauma after lens implantation.

PURPOSE: To examine the ultrastructure of clear corneal incisions (CCIs) performed with diamond keratomes and steel blades as well as the corneal trauma after implantation of a foldable intraocular lens (IOL) through two incision widths. SETTING: University Eye Clinic and Institute of Histology and Embryology II, University of Vienna, Austria. METHODS: Twenty-four human cadaver eyes without prior ocular surgery were obtained from the University Eye Bank, Vienna. Single-plane CCIs were performed with 3.0 and 3.2 mm Alcon steel blades and with a 3.0 mm Huco diamond keratome. The AMO PhacoFlex II lens was implanted with a Fine II folder. During the entire procedure, the eye pressure was kept between 26 and 30 mm Hg by infusing balanced salt solution into the anterior chamber. Specimens were prepared for light microscopy, transmission electron microscopy, and scanning electron microscopy according to standard procedures. RESULTS: The diamond keratome produced cleaner cuts than the steel blade. After IOL implantation, 3.0 mm steel blade incisions exhibited extensions at their lateral ends. Within these extensions, the collagen lamellae were displaced and torn. This was not true with 3.2 mm tunnels. Because of the thickness of a 3.0 mm diamond keratome, the extent of corneal trauma was between that found with 3.0 and 3.2 mm steel keratome tunnels. CONCLUSIONS: Implantation of the SI-30 through 3.0 mm CCIs produced by the steel blade led to more severe corneal trauma than implantation through 3.2 mm steel blade incisions or 3.0 mm diamond keratome incisions. Thus, IOL implantation through incisions that are too small intensifies corneal trauma.