VEGF enhances functional improvement of postinfarcted hearts by transplantation of ESC-differentiated cells.

Despite considerable advances in medicine, the incidence of heart failure remains high in patients after myocardial infarction (MI). This study investigated the effects of engrafted early-differentiated cells (EDCs) from mouse embryonic stem cells, with or without transfection of vascular endothelial growth factor (VEGF) cDNA (phVEGF(165)), on cardiac function in postinfarcted mice. EDCs were transfected with green fluorescent protein (GFP) cDNA and transplanted into infarcted myocardium. Compared with the MI mice receiving cell-free medium, cardiac function was significantly improved in the MI mice 6 wk after transplantation of EDCs. Moreover, improvement of heart function was significantly greater in the mice implanted with EDCs overexpressing VEGF (EDCs-VEGF) than with EDCs alone. Frozen sections of infarcted myocardium with EDCs or EDCs-VEGF transplantation showed GFP-positive tissue. The area with positive immunostaining for cardiac troponin I and alpha-myosin heavy chain was larger in injured myocardium with EDCs or EDCs-VEGF transplantation than with medium injection. Transplantation of EDCs or EDCs-VEGF significantly increased the number of blood vessels in the MI area. However, the density of capillaries was significantly higher in the EDCs-VEGF animals than in the EDC mice. Double staining for GFP and connexin-43 was positive in injured myocardium with EDC transplantation. Our data demonstrate that engrafted EDCs or EDCs-VEGF regenerated cardiac tissue and significantly improved cardiac function in postinfarcted hearts. The novel EDCs-VEGF synergistic approach may have an important impact on future cell therapy for patients experiencing MI or heart failure.     

Is cryopreservation a homogeneous process? Ultrastructure and motility of untreated, prefreezing, and postthawed spermatozoa of Diplodus puntazzo (Cetti).

This study subdivides the cryopreservation procedure for Diplodus puntazzo spermatozoa into three key phases, fresh, prefreezing (samples equilibrated in cryosolutions), and postthawed stages, and examines the ultrastructural anomalies and motility profiles of spermatozoa in each stage, with different cryodiluents. Two simple cryosolutions were evaluated: 0.17 M sodium chloride containing a final concentration of 15% dimethyl sulfoxide (Me(2)SO) (cryosolution A) and 0.1 M sodium citrate containing a final concentration of 10% Me(2)SO (cryosolution B). Ultrastructural anomalies of the plasmatic and nuclear membranes of the sperm head were common and the severity of the cryoinjury differed significantly between the pre- and the postfreezing phases and between the two cryosolutions. In spermatozoa diluted with cryosolution A, during the prefreezing phase, the plasmalemma of 61% of the cells was absent or damaged compared with 24% in the fresh sample (P < 0.001). In spermatozoa diluted with cryosolution B, there was a pronounced increase in the number of cells lacking the head plasmatic membrane from the prefreezing to the postthawed stages (from 32 to 52%, P < 0.01). In both cryosolutions, damages to nuclear membrane were significantly higher after freezing (cryosolution A: 8 to 23%, P < 0.01; cryosolution B: 5 to 38%, P < 0.001). With cryosolution A, the after-activation motility profile confirmed a consistent drop from fresh at the prefreezing stage, whereas freezing and thawing did not affect the motility much further and 50% of the cells were immotile by 60-90 s after activation. With cryosolution B, only the postthawing stage showed a sharp drop of motility profile. This study suggests that the different phases of the cryoprocess should be investigated to better understand the process of sperm damage.     

A significant increase in both basal and maximal calcineurin activity in the rat pilocarpine model of status epilepticus

This study focused on the effects of status epilepticus on the activity of calcineurin, a neuronally enriched, calcium-dependent phosphatase. Calcineurin is an important modulator of many neuronal processes, including learning and memory, induction of apoptosis, receptor function and neuronal excitability. Therefore, a status epilepticus-induced alteration of the activity of this important phosphatase would have significant physiological implications. Status epilepticus was induced by pilocarpine injection and allowed to continue for 60 min. Brain region homogenates were then assayed for calcineurin activity by dephosphorylation of p-nitrophenol phosphate. A significant status epilepticus-dependent increase in both basal and Mn(2+)-dependent calcineurin activity was observed in homogenates isolated from the cortex and hippocampus, but not the cerebellum. This increase was resistant to 150 nM okadaic acid, but sensitive to 50 microM okadaic acid. The increase in basal activity was also resistant to 100 microM sodium orthovanadate. Both maximal dephosphorylation rate and substrate affinity were increased following status epilepticus. However, the increase in calcineurin activity was not found to be due to an increase in calcineurin enzyme levels. Finally, increase in calcineurin activity was found to be NMDA-receptor activation dependent. The data demonstrate that status epilepticus resulted in a significant increase in both basal and maximal calcineurin activity.     

Functional anatomy of five endangered tropical timber wood species of the family Dipterocarpaceae

Wood anatomy of five dipterocarp species endemic to the Philippines was studied with the goal to explore functional wood traits of ecological significance. Stem wood of 6-year-old trees grown under similar environmental conditions in a plantation (Leyte, Philippines) was used. Wood densities decreased in the following order Hopea plagata > Dipterocarpus kerrii > Parashorea malaanoman > Shorea almon ≈ Shorea contorta. This was mainly caused by significantly thicker fiber cell walls of H. plagata and D. kerrii than those of the other three species. Wood density was negatively correlated with the abundance of axial parenchyma cells. Predicted conductance was independent from wood density and lowest in H. plataga and highest in D. kerrii and S. contorta. These results indicate that H. plagata and D. kerrii is woods have higher constructions costs in term of carbon per unit of biomass, and that H. plagata is probably better acclimated to varying soil moisture than the other species.     

Regulation of glutamate dehydrogenase activity by amino acids in maize seedlings

Root or secondary leaf segments from maize ( Zea mays L. cv. Ganga safed-2) seedlings were incubated with 9-amino acids and two amides separately, each at 5 m M for 24 h, to study their effects on glutamate dehydrogenase (GDH) activity. Most of the compounds tested inhibited the specific activity of NADH-GDH and increased that of NAD⁺-GDH in the roots in the presence as well as in the absence of ammonium. In the leaves, such effects were recorded only with a few amino acids. Total soluble protein in the root and leaf tissues increased with the supply of most of the amino compounds. The effect of glutamate on enzyme activity and protein was concentration dependent in both tissues. When the enzyme extracts from root or leaf tissues were incubated with some of the amino acids, NADH-GDH declined while NAD⁺-GDH increased in most cases. The inhibition of NADH-GDH increased with increasing concentration of cysteine from 1 to 5 m M . The experiments demonstrate that most of the amino acids regulated GDH activity, possibly through some physicochemical modulation of the enzyme molecule.     

Small RNAs: regulators and guardians of the genome

Small non-coding RNAs comprise several classes and sizes, but all share a unifying function in cellular physiology: epigenetic regulation of gene expression. Here, we review the salient aspects of recent studies on the biogenesis and function of three classes of small RNAs: miRNAs, siRNAs, and piRNAs. Although the mechanisms are becoming clear by which siRNA-triggered mRNA cleavage silences genes, more studies are needed on several issues regarding miRNA-mediated translation repression. Piwi proteins have been suggested to co-operate in amplifying piRNA biogenesis to maintain transposon silencing in the germ line genome, but details of this process are still unknown as well as the functional consequences of piRNA expression at discrete genomic loci.