Patients with urological disorders may benefit from gene based therapy. We investigated the feasibility of delivering exogenous genes into urological tissues in vivo using direct in vivo electrotransfection.Gene transfer to rat kidneys, testes and bladders was accomplished via direct local injection of pGL3/luciferase and beta-galactosidase reporter gene constructs, followed by an electrical pulse ranging from 55 to 115 msec at 100 V. Direct injection of deoxyribonucleic acid without an electrical pulse served as the control. The transfected and nontransfected organs were retrieved and analyzed by luciferase activity assay, histochemical and immunocytochemical staining for beta-galactosidase, and reverse transcription polymerase chain reaction with primers specific for beta-galactosidase messenger ribonucleic acid.There was significant luciferase activity 1, 3 and 5 days after direct in vivo electrotransfection in kidneys and testes, and after 3, 5, 7 and 10 days in bladders. Positive beta-galactosidase enzyme activity and beta-galactosidase immunoreactivity were observed in the transfected renal tubular cells, testicular interstitial and germ cells, and uroepithelial bladder layer. Reverse transcription-polymerase chain reaction products of the transfected organs were noted, indicating the successful transcription of messenger ribonucleic acid.This study demonstrates that direct in vivo electrotransfection is a feasible method of transient gene delivery into intact urological organs. Its apparent safety and relative simplicity suggest that direct in vivo electrotransfection may be useful clinically.
The sensitivity of the total ozone distribution in our two‐dimensional photochemical model to dynamical inputs has been explored. The residual circulation has been computed from three sets of heating rates but using the same temperature distributions. These heating rates result in advection fields that are appropriately linked with the magnitude of their vertical winds in the lower stratosphere and are called: (1) the strong circulation, (2) the weak circulation, and (3) the combined circulation. Three different formulations of horizontal eddy diffusion have also been investigated, including (1) a small constant diffusion, (2) a computed diffusion from potential vorticity using measured temperatures, and (3) a self‐consistent diffusion determined from the meridional velocity. Finally, we studied two different formulations of vertical eddy diffusion: (1) a small constant diffusion and (2) an altitude‐dependent larger diffusion. Our modeled global average total ozone varies by up to 10%, depending on the various dynamical inputs, and the seasonal and latitudinal variabilities are even more substantial. The calculations indicate that total ozone in the middle to high latitudes is very sensitive to the advection field below 100 mbar, the region where the heating rates (which are used in computing the advection field) are most uncertain. Modeled total ozone shows better agreement with solar backscattered ultraviolet (SBUV) ozone climatology, when computed horizontal eddy diffusion is used. Our “best” modeled total ozone distribution is a result of using the heating rates from Rosenfield et al. (1987) for pressures less than 100 mbar, along with heating rates from Dopplick (1974, 1979) for pressures greater than 100 mbar. Our model results, especially when comparing model and SBUV ozone between 10 and 1 mbar, indicate that there is an inconsistency between our “best” horizontal eddy diffusion and the “best” mean residual circulation. Our studies indicate a relative similarity of a typical ozone perturbation scenario among the different dynamical inputs. All of the perturbation studies predict the smallest ozone changes in the tropical latitudes, with larger ozone changes in the middle to high latitudes. The major changes in the various perturbation results are in the middle to high latitudes: (1) the peak ozone change is off of the pole in the southern hemisphere (near 60°S latitude) using the strong circulation but centered on the pole for the various other dynamical studies, and (2) the maximum ozone change is centered at times varying from early spring to early summer, depending on which dynamical input is used.
The objective of this research was to examine the effects of chemical perturbations on the stratosphere using models which can incorporate fully interactive radiative, chemical, and dynamical responses, in the context of a zonally averaged model. Model runs for the unperturbed, chlorine-perturbed and simultaneously chlorine-and CO2-perturbed cases were completed using the JPL-87 chemical kinetics data. The base case was analyzed and submitted for publication. The perturbed cases show substantial sensitivity of the predicted column ozone depletion to the perturbations affecting lower stratosphere temperature, but less to far dynamical perturbations. The column ozone distribution changed substantially when the kinetics data was changed. This implies a greater-than-expected uncertainty in predicted latitude distributions of ozone depletion, due to uncertainty about the accuracy and completeness of the chemical kinetics data set.
Proglumide, a glutaramic acid derivative, inhibits the acid secretory effects of gastrin and is said to be a specific gastrin receptor blocking agent. In the current study we have shown that proglumide inhibits the binding of gastrin to its receptor in rat oxyntic gland mucosa and tested whether this receptor is also responsible for the trophic effect of gastrin. In the first study 400 mg/kg proglumide injected with 250 micrograms/kg pentagastrin every 8 h for 48 h totally prevented the trophic effects of pentagastrin in rat oxyntic gland mucosa. Parameters measured included DNA synthesis and DNA, RNA, and protein content. In a second study the lowest maximally effective dose of proglumide was determined to be 100 mg/kg. The trophic effect of pentagastrin was inhibited in duodenal mucosa, colonic mucosa, and pancreas as well as oxyntic gland mucosa. These data demonstrate that proglumide blocks the trophic action of exogenous gastrin and suggest that the trophic effect of gastrin is mediated by the gastrin receptor.
The sensitivity of the N 2 O distribution in our two‐dimensional photochemical model to horizontal diffusion coefficients (both K yy and K yz values) has been explored. The residual circulation was computed using temperature data from the National Meteorological Center (NMC) and heating rates from Rosenfield et al. (1987). The K yy and K yz values were computed from these same data using quasigeostrophic potential vorticity. For comparison we also computed self‐consistent K yy values from our residual circulation. The use of either set of these diffusion coefficients produced substantial changes in the N 2 O distribution, especially in the middle to upper stratosphere and at high latitudes in the winter. In general, the changes lead to an enhanced transport of N 2 O to higher latitudes with the set of self‐consistent K yy values transporting the greatest amount of N 2 O. The enhanced transport increases the lifetime of N 2 O because the photochemical lifetime is larger at high latitudes.
Spontaneous transformation in continuous culture of the androgen-sensitive rat prostate fibroblast cell line, NbF-1, resulted in an aggressively tumorigenic nonmetastatic phenotype that coincided with few gross chromosome abnormalities. This study identified transformation-associated alterations in extracellular matrix and androgen receptor expression in the NbF-1 cell line. Substantial levels of procollagens I, III, and IV and fibronectin mRNAs were detected in nontumorigenic NbF-1 cells. Laminin B1 and B2 mRNAs were also detectable, but at lower levels. Expression of all six extracellular matrix mRNAs was nonuniformly lower in tumorigenic NbF-1 cells. This decrease in expression was greatest for alpha 2 procollagen IV mRNA, which was reduced 17-fold. Proteoglycans and glycosaminoglycans synthesized by the NbF-1 cultures were also characterized. The NbF-1 cell line expressed chondroitin sulfate proteoglycans predominantly, and expression was reduced 5- to 10-fold in tumorigenic cultures. In contrast to the extensive alterations in the extracellular matrix, measurement of high-affinity androgen binding and androgen receptor mRNA levels showed substantial expression of androgen receptors in both NbF-1 cultures. Cultures of early and late passage NbF-1 cells demonstrated a mitogenic response to dihydrotestosterone. These data indicate (a) that alterations in expression of extracellular matrix components may represent early markers for tumorigenic transformation in prostatic mesenchymal cells and (b) that these changes can occur without disrupting androgen receptor expression and androgen sensitivity.
Several factors contribute to the errors encountered. With the exception of the line-by-line model, all of the models employ simplifying assumptions that place fundamental limits on their accuracy and range of validity. For example, all 2-D modeling groups use the diffusivity factor approximation. This approximation produces little error in tropospheric H2O and CO2 cooling rates, but can produce significant errors in CO2 and O3 cooling rates at the stratopause. All models suffer from fundamental uncertainties in shapes and strengths of spectral lines. Thermal flux algorithms being used in 2-D tracer tranport models produce cooling rates that differ by as much as 40 percent for the same input model atmosphere. Disagreements of this magnitude are important since the thermal cooling rates must be subtracted from the almost-equal solar heating rates to derive the net radiative heating rates and the 2-D model diabatic circulation. For much of the annual cycle, the net radiative heating rates are comparable in magnitude to the cooling rate differences described. Many of the models underestimate the cooling rates in the middle and lower stratosphere. The consequences of these errors for the net heating rates and the diabatic circulation will depend on their meridional structure, which was not tested here. Other models underestimate the cooling near 1 mbar. Suchs errors pose potential problems for future interactive ozone assessment studies, since they could produce artificially-high temperatures and increased O3 destruction at these levels. These concerns suggest that a great deal of work is needed to improve the performance of thermal cooling rate algorithms used in the 2-D tracer transport models.
