Long term peripheral nerve regeneration using a novel PCL nerve conduit.

The gold standard in surgical management of a peripheral nerve gap is currently autologous nerve grafting. This confers patient morbidity and increases surgical time therefore innovative experimental strategies towards engineering a synthetic nerve conduit are welcome. We have developed a novel synthetic conduit made of poly ε-caprolactone (PCL) that has demonstrated promising peripheral nerve regeneration in short-term studies. This material has been engineered to permit translation into clinical practice and here we demonstrate that histological outcomes in a long-term in vivo experiment are comparable with that of autologous nerve grafting. A 1cm nerve gap in a rat sciatic nerve injury model was repaired with a PCL nerve conduit or an autologous nerve graft. At 18 weeks post surgical repair, there was a similar volume of regenerating axons within the nerve autograft and PCL conduit repair groups, and similar numbers of myelinated axons in the distal stump of both groups. Furthermore, there was evidence of comparable re-innervation of end organ muscle and skin with the only significant difference the lower wet weight of the muscle from the PCL conduit nerve repair group. This study stimulates further work on the potential use of this synthetic biodegradable PCL nerve conduit in a clinical setting.

Retention of the developmental pluripotency in medaka embryonic stem cells after gene transfer and long-term drug selection for gene targeting in fish

Embryonic stem (ES) cells provide a unique tool for introducing random or targeted genetic alterations, because it is possible that the desired, but extremely rare recombinant genotypes can be screened by drug selection. ES cell-mediated transgenesis has so far been limited to the mouse. In the fish medaka (Oryzias latipes) several ES cell lines have been made available. Here we report the optimized conditions for gene transfer and drug selection in the medaka ES cell line MES1 as a prelude for gene targeting in fish. MES1 cells gave rise to a moderate to high transfection efficiency by the calcium phosphate co-precipitation (5%), commercial reagents Fugene (11%), GeneJuice (21%) and electroporation (>30%). Transient gene transfer and CAT reporter assay revealed that several enhancers/promoters and their combinations including CMV, RSV and ST (the SV40 virus early gene enhancer linked to the thymidine kinase promoter) were suitable regulatory sequences to drive transgene expression in the MES1 cells. We show that neo, hyg or pac conferred resistance to G418, hygromycin or puromycin for positive selection, while the HSV-tk generated sensitivity to ganciclovir for negative selection. The positive-negative selection procedure that is widely used for gene targeting in mouse ES cells was found to be effective also in MES1 cells. Importantly, we demonstrate that MES1 cells after gene transfer and long-term drug selection retained the developmental pluripotency, as they were able to undergo induced differentiation in vitro and to contribute to various tissues and organs during chimeric embryogenesis.

Long-term (1956-1999) dynamics of phosphorus in a shallow, subtropical Chinese lake with the possible effects of cyanobacterial blooms

This paper describes the long-term dynamics of phosphorus concentrations in both the lake water and the sediment in a subtropical Chinese lake, Lake Donghu. The total phosphorus (TP) concentration in the lake water experienced an upward trend from the 1950s, and peaked in 1983/1984, but declined obviously afterwards. From the 1950s to the 1990s, TP content in the upper 10 cm sediment of the lake increased steadily from 0.307 to 1.68 mg Pg DW-1 at Station I and from 0.151 to 0.89 mg Pg DW-1 at Station II, respectively. The TP increase in the lake water before mid-1980s was mainly attributed to the massive input of sewage P. The outbreak of cyanobacterial blooms coincided with the peaks of TP and Orthophosphate (PO4-P) in the water in mid-1980s, and the maximum TP of the water reached as high as 1.349 mg/1 at Station I and 0.757 mg/l at Station II (in 1984), respectively. The declines of TP and PO4-P in the water after mid-1980s was coincident with the disappearance of cyanobacterial bloom. (C) 2001 Elsevier Science Ltd. All rights reserved.

Long-term changes of planktonic rotifers in a subtropical Chinese lake dominated by filter-feeding fishes

1. The long-term changes (1956-1998) in density and species composition of planktonic rotifers were studied at two sampling stations (I, II) of Lake Donghu, a shallow eutrophic Chinese Lake densely stocked with filter-feeding fishes. Annual average densities of rotifers increased with an increase in fish yield and eutrophication, whilst species number decreased from 82 in 1962-1963 to 62 in 1994-1998. 2. During 1962-98, some species such as Anuraeopsis fissa, Polyarthra spp. (including P. dolichoptera & P. vulgaris), Trichocerca pusilla and Synchaeta oblonga increased their percentage in abundance remarkably, whilst the proportion of Keratella cochlearis decreased at two relatively eutrophic stations from 19 to 4.2% at Station I and from 30 to 3.2% at Station IL 3. The high r(max) of A. fissa probably made it more successful than other rotifers under high predation pressure by planktivorous fish. The decrease in the K. cochlearis population might be attributed partly to predation by Cyclops vicinus. 4. Small rotifers were less vulnerable to fish predation than large-sized cladocerans. Decreases in cladocerans coincided with increases in rotifers, suggesting that the indirect effect of fish predation on cladocerans might have partly contributed to the population development of rotifers in Lake Donghu during recent decades. 5. We also conducted surveys (1994-1998) of seasonal dynamics of rotifers at four sampling stations (I-IV) which have varied in trophic status after fragmentation of the lake in the 1960s. A total of 75 species were identified at the four stations. Both densities and biomass of rotifers were considerably higher in the two more eutrophic stations than in the two less eutrophic stations. This indicates that the population increase of rotifers at Stations I and II during recent decades might be partly attributed to eutrophication of the lake water.

Impacts of filter-feeding fishes on the long-term changes of crustacean zooplankton in a eutrophic subtropical Chinese lake

Long-term changes In the crustacean zooplankton community (calanoid and cyclopoid copepods and cladocerans) were studied in Lake Donghu, a shallow and eutrophic Chinese lake. This lake had been earlier stocked with two pump Alter-feeding Ashes, silver carp (Hypopthalmichthys molitrix) and bighead carp (Aristichthys nobilis). During the 1950s and the mid-1980s, the ratio of copepods to cladocerans was relatively stable but showed a general increase thereafter. From the early-1980s to the 1990s, calanoid/cyclopoid ratios decreased obviously. In the 1990s, Cyclops vicinus, Diaphanosoma brachyurum, and Moina micrura were dominant the abundance of C. vicinus and M. micrura increased significantly; and D, brachyurum showed a substantial decrease. The study shows that under extremely high pressure of Ash predation, the species which could recover rapidly from fish predation would be the most likely to survive and increase their numbers.

Long-term toxic impact of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the reproduction, sexual differentiation, and development of different life stages of Gobiocypris rarus and Daphnia magna

The sexual ratio of Gobiocypris rarus exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and 17 beta -estradiol from embryo to sexually mature revealed feminization and overdevelopment of connective tissue in male fish gonad in 2-30 pg/L TCDD concentration range. Daphnia magna was not sensitive to the high dose of TCDD (0.1-1000 ng/ml), but the reproduction of D. magna treated with TCDD decreased after the 8th day. 7-Ethoxyresorufin-O-deethylase (EROD) activities in newly fertilized eggs of G. rarus exposed to TCDD dosage groups (1000-100,000 pg/L) were significantly induced and increased with TCDD concentrations at the early life stage, while no difference was found between low TCDD dosage groups (<100 pg/L), but a good relationship between the EROD activity and the TCDD concentration was observed during a long-term developmental stage. There was a pericardial edema formed in a 2-week yolk-sac at the concentration of 1000 pg/L TCDD. But in the exposure group (2 pg/L TCDD for 120 days), the cell nuclei of hepatocytes was far from the center and packed toward the cell membrane; the cristae of most mitochondria in the cell dropped and collapsed; the rough endoplasmic reticulum broke into fragments; and numerous lipid droplets formed in the cell. (C) 2001 Academic Press.

Long-term changes of Copepoda community (1957-1996) in a subtropical Chinese lake stocked densely with planktivorous filter-feeding silver and bighead carp

In contrast to the relatively well documented impact of particulate-feeding fish on zooplankton communities, little attention has been devoted to the impact of filter-feeding fish. Filter-feeding silver and bighead carp are the most intensively cultured fish species in Asia and comprise much of the production of Chinese aquaculture. However, little information is known about the impact of either fish on the zooplankton community. Long-term changes in the Copepoda community (1957-1996) were studied at two sampling stations of a subtropical Chinese lake (Lake Donghu) dominated by silver and bighead carp. For both calanoids and cyclopoids, the littoral station (I) was much more resource profitable than the pelagic station (II). There has been a tremendous increase in the annual fish catch over the past 30 years due to the increased stocking with fingerlings of the two carp species. There was a notably higher fish density at Station I than at Station II. Cyclopoid abundance was notably higher at Station I than at Station II during the 1950s to the 1980s, while the reverse became true in the 1990s. This is probably because when fish abundance increased to an extremely high level, the impact of fish predation on the cyclopoids became more important than that of food resources at the littoral station. At both stations, cyclopoid abundance was relatively low in spite of the presence of abundant prey. Similarly, calanoid density did not differ significantly between the two stations in the 1950s and 1960s, but was significantly lower at Station I than at Station II during the 1980s and 1990s. Such changes are attributed to the gradient of fish predation between the stations and an increasing predation pressure by the fish. The increased fish predation also correlated with a shift in summer-dominant calanoids from larger species to smaller ones. In conclusion, the predaceous cyclopoids are affected by fish predation to a much lesser extent than the herbivorous calanoids, and therefore increased predation by filter-feeding fish results in a definite increase in the cyclopoid/calanoid ratio. Predation by filter-feeding fish has been a driving force in shaping the copepod community structure of Lake Donghu during the past decades.

Distribution, transformation, and long-term accumulation of polychlorinated dibenzo-p-dioxins and dibenzofurans in different tissues of fish and piscivorous birds

The present study monitored 10-year-old fish and piscivorous birds from sites contaminated for many Stars. The data reflected the results of actual, long-term environmental exposures, The results demonstrate that different tissues of fish have quite different concentrations of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/F), The concentration order of PCDD/F within fish is liver congruent to egg congruent to intestines kidney congruent to hearts gill congruent to bladders > muscle > brain. The concentration order of PCDD/F within piscivorous birds was livers egg congruent to hearts muscle congruent to stomachs brain, The results obtained also demonstrate that the accumulation patterns of piscivorous birds and fish are quite different. The tissues of fish and piscivorous birds have different capacities for bioaccumulation and biotransformation of PCDD/F; variable proportions of TEQs were also found throughout their bodies. In fish, toxic equivalency quotient (TEQ): PCDD/F ratios in various tissues ranged from 0.01 to 0.07, whereas in birds the ratios ranged from 0.07 to 0.43. If the concentrations are normalized with lipid content, the results vary less. The effect of different lipid properties is obvious in the case of brain tissue, which is richer in phospholipids. (C) 2000 Academic Press.

Long-term changes in crustacean zooplankton and water quality in a shallow, eutrophic Chinese lake densely stocked with fish

From surveys made in 1962-1963, 1973-1974, 1979-1996 at two Stations in Lake Donghu, a shallow eutrophic water body near Wuhan, P. R. China, the authors, derive long-term changes in species composition, standing crop and body-size of planktonic crustaceans. The species number decreased from the 1960s to the 1990s. The cladocerans dropped from 46 (1960s) to 26 (1980s) to 13 (1990s); the copepods decreased from 14 (1960s) to 10 (1980s) to 7 (1990s). From the mid-1980s on, the dominant crustaceans also changed: Daphnia hyalina and D. carinata ssp. were replaced by Moina micrura and Diaphanosoma brachyurum at Stations 1 and 2, respectively; Cyclops vicinus replaced Mesocyclops leuckarti. Densities and biomass of Cladocera decreased markedly after 1987. Annual average densities and biomass of cladocerans were statistically differences between 1962-1986 and 1987-1996 (P < 0.01). Annual average densities of Daphnia (Station 1 + Station 2) were negatively correlated with fish yield. Since the 1980s, annual average body length of Cladocera and Calanoida decreased, while annual average body length of Cyclopoida increased. In the same years, average body length of copepods was lower during May-October than during January-April and November-December. A 12-yr data analysis showed annual average concentration of chlorophyll-a (Chl-a) to be negatively correlated with annual average density of Daphnia, whilst lake transparency was positively correlated with annual average densities of Daphnia. The results imply that, since Daphnia feeds efficiently on phytoplankton, it could decrease concentration of Chl-a, and enhance water transparency.

STUDY OF THE LONG-TERM STABILITY OF THE EFFECTIVE CONCENTRATION OF IONIZED SPACE CHARGES (N-EFF) OF NEUTRON-IRRADIATED SILICON DETECTORS FABRICATED BY VARIOUS THERMAL-OXIDATION PROCESSES

Experimental study of the reverse annealing of the effective concentration of ionized space charges (N-eff, also called effective doping or impurity concentration) of neutron irradiated high resistivity silicon detectors fabricated on wafers with various thermal oxides has been conducted at room temperature (RT) and elevated temperature (ET). Various thermal oxidations with temperatures ranging from 975 degrees C to 1200 degrees C with and without trichlorethane (TCA), which result in different concentrations of oxygen and carbon impurities, have been used. It has been found that, the RT annealing of the N-eff is hindered initially (t < 42 days after the radiation) for detectors made on the oxides with high carbon concentrations, and there was no carbon effect on the long term (t > 42 days after the radiation) N-eff reverse annealing. No apparent effect of oxygen on the stability of N-eff has been observed at RT. At elevated temperature (80 degrees C), no significant difference in annealing behavior has been found for detectors fabricated on silicon wafers with various thermal oxides. It is apparent that for the initial stages (first and/or second) of N-eff reverse annealing, there may tie no dependence on the oxygen and carbon concentrations in the ranges studied.

The development of long - Term ambulatory pH monitoring system and its clinical application

This paper introduced a long-term ambulatory intragastric pH monitoring system, which is designed for prolonged ambulatory studies of Gastroesophageal Reflux Diseases. The whole system is composed of the gastric catheter with two pH sensors, a small data logger (Microdatalog), and a notebook PC. In this paper, the design of monitoring system hardware and software are described in detail. Clinical applications reveal good results.