Application of Stereolithographic Custom Models for Studying the Impact of Biofilms and Mineral Precipitation on Fluid Flow

Here we introduce the use of transparent experimental models fabricated by stereolithography for studying the impacts of biomass accumulation, minerals precipitation, and physical configuration of flow paths on liquid flow in fracture apertures. The internal configuration of the models ranged in complexity from simple geometric shapes to those that incorporate replicated surfaces of natural fractures and computationally derived fracture surfaces. High-resolution digital time-lapse imaging was employed to qualitatively observe the migration of colloidal and soluble dyes through the flow models. In this study, a Sphingomonas sp. and Sporosarcina (Bacillus) pasteurii influenced the fluid dynamics by physically altering flow paths. Microbial colonization and calcite deposition enhanced the stagnant regions adjacent to solid boundaries. Microbial growth and calcite precipitation occurred to a greater extent in areas behind the fabricated obstacles and less in high-velocity orifices.     

Blood flow response to a postural challenge in older men and women

BACKGROUND: The purpose of this study was to measure blood flow in the carotid and femoral arteries, heart rate and blood pressure in response to postural challenge in older adults. A second purpose was to determine if older men and women have different cardiovascular responses to a postural challenge such as tilt. METHODS: Thirty-seven healthy elderly men and women participated in this study (69-82 years old). All subjects had similar physical activity levels. Postural challenge was induced by a 60 degrees tilt at the level of the waist. Continuous carotid blood flow and femoral blood flow was measured with Doppler ultrasound. RESULTS: Carotid blood flow was significantly reduced 17% in both men and women immediately after tilt (p < 0.001), and by 3.2% two minutes after tilt (p < 0.001). Femoral blood flow decreased 59.4% in men and 61% in women immediately after tilt (p < 0.001), and remained significantly decreased two minutes after tilt by 21% (p <0.001). Heart rate increased by 15% in men (p < 0.001), and 26% in women immediately after the tilt (p < 0.001). Heart rate returned to resting values within two minutes in both men and women. Response to tilt was not significantly related to self-report physical activity levels or to six-minute walk time. CONCLUSION: A postural challenge induced larger changes in the femoral artery compared to the carotid artery. There were no differences between men and women to a tilt table test except for differences in heart rate response. There was no difference in the blood flow responses to postural challenge with physical activity level or between healthy older men and women.     

Crosstalk between estrogen receptor alpha and the aryl hydrocarbon receptor in breast cancer cells involves unidirectional activation of proteasomes

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is an environmental toxin that activates the aryl hydrocarbon receptor (AhR) and disrupts multiple endocrine signaling pathways. T47D human breast cancer cells express a functional estrogen receptor alpha (ERalpha) and AhR, and treatment of these cells with 17beta-estradiol (E2) or TCDD resulted in a rapid proteasome-dependent decrease in immunoreactive ERalpha and AhR proteins (>60-80%), respectively. E2 did not affect the AhR, whereas TCDD induced proteasome-dependent degradation of both the AhR and ERalpha in T47D and MCF-7 human breast cancer cells, and these responses were specifically blocked by proteasome inhibitors. Thus, TCDD-induced degradation of ERalpha may contribute to the antiestrogenic activity of AhR agonists and this pathway may be involved in AhR-mediated disruption of other endocrine responses.     

Comparative use of color vision for frugivory by sympatric species of platyrrhines

Ateles spp. and Alouatta spp. are often sympatric, and although they are mainly frugivorous and folivorous, respectively, they consume some of the same fruit species. However, they differ in terms of color vision, which is thought to be important for fruit detection. Alouatta spp. have routine trichromatic color vision, while Ateles spp. presents the classic polymorphism of platyrrhines: heterozygous females have trichromatic color vision, and males and homozygous females have dichromatic vision. Given these perceptual differences, one might expect Alouatta spp. to consume more reddish fruits than Ateles spp., since trichromats have an advantage for detecting fruits of that hue. Furthermore, since Ateles spp. have up to six different color vision phenotypes, as do most other platyrrhines, they might be expected to include fruits with a wider variety of hues in their diet than Alouatta spp. To test these hypotheses we studied the fruit foraging behavior of sympatric Alouatta palliata and Ateles geoffroyi in Costa Rica, and modeled the detectability of fruit via the various color vision phenotypes in these primates. We found little similarity in fruit diet between these two species (Morisita = 0.086). Furthermore, despite its polymorphism, A. geoffroyi consumed more reddish fruits than A. palliata, which consumed more greenish fruits. Our modeling results suggest that most fruit species included in the diet of A. geoffroyi can be discriminated by most color vision phenotypes present in the population. These findings show that the effect of polymorphism in platyrrhines on fruit detection may not be a disadvantage for frugivory. We suggest that routine trichromacy may be advantageous for other foraging tasks, such as feeding on young leaves.     

Substrate specificity of rat brain calcium-activated and phospholipid-dependent protein kinase

A synthetic peptide ArgThrProProProSerGly with sequence similar to the threonine sites of phosphorylation in both myelin basic protein and simian virus 40 T antigen could be phosphorylated in vitro by a purified rat brain Ca2+-activated and phospholipid-dependent protein kinase, protein kinase C. The apparent Km and Vm values of this heptapeptide for the enzyme were determined to be 240 microM and 60 nmol/min/mg, respectively. Up to 0.8 mol 32P could be incorporated into the peptide, mainly at the threonine residue. Substitution of the L-threonine residue in the heptapeptide by its D-enantiomer abolished the phosphorylatability of the peptide by protein kinase C. However, this (D)Thr-containing peptide could act as a competitive inhibitor for the kinase with an apparent Ki value of approximately 320 microM. These findings suggest that a triprolyl sequence may act as a recognition site for protein kinase C.     

Regulation of proliferation and differentiation of respiratory tract epithelial cells by TGF beta

In this paper we examined the effects of transforming growth factor beta (TGF beta) on the proliferation and differentiation of rabbit tracheal epithelial cells in primary culture. Treatment of these cells with TGF beta inhibits cell proliferation in a time- and dose-dependent manner; concentrations as low as 1 pM are able to inhibit cell growth. Concomitantly, TGF beta causes cells to accumulate in the G0/G1 phase of the cell cycle and a sharp reduction in the ability of the cells to form colonies after subculture at clonal density. These results indicate that TGF beta induces terminal cell division in these cells. The inhibition of cell growth is accompanied by changes in cell morphology and a stimulation of the formation of cross-linked envelopes. TGF beta enhances the levels of transglutaminase activity and cholesterol sulfate, two markers of squamous differentiation. Our results indicate that TGF beta induces terminal squamous cell differentiation in rabbit tracheal epithelial cells. Retinoic acid (RA) does not affect the commitment to terminal cell division induced by TGF beta, but inhibits the expression of the squamous phenotype. Growth of normal human bronchial epithelial cells was affected by TGF beta in a way similar to that of rabbit tracheal epithelial cells. Several carcinoma cell lines tested were quite resistant to TGF beta, whereas growth of one carcinoma cell line was stimulated by TGF beta. These results indicate that a modified response to TGF beta could be one mechanism involved in the aberrant growth control of malignant cells.     

Comparative properties of untreated andN-nitrosobenzylmethyl-amine-transformed rat esophageal epithelial cell lines

Summary A culture system utilizing rat esophageal epithelial cells has been developed. Four normal and eightN-nitrosobenzylmethylamine-treated lines were compared with respect to chromosome number, anchorage-independent growth in agarose, and tumorigenic potential in syngenic rats. All cell lines were aneuploid with nine in the near-tetraploid range and three in the near-diploid range. No relation between tumorigenic potential and chromosome number or structure was apparent. Similarly, anchorage-independent growth in agarose did not correlate with tumorigenic potential. Three of the 12 immortalized lines (two carcinogen-treated and 1 untreated) induced well-differentiated squamous cell carcinomas in syngeneic rats. These tumors had weak metastatic potentials suggesting that tumorigenic potential and metastatic ability are separately controlled. These cell lines will be useful for the investigation of factors involved in the conversion of immortalized rat esophageal epithelial cell lines to lines of high metastatic potential.     

Comparison of methods to measure acute metal and organometal toxicity to natural aquatic microbial communities.

Microbial communities in water from Baltimore Harbor and from the mainstem of Chesapeake Bay were examined for sensitivity to mercuric chloride, monomethyl mercury, stannic chloride, and tributyltin chloride. Acute toxicity was determined by measuring the effects of [3H]thymidine incorporation, [14C]glutamate incorporation and respiration, and viability as compared with those of controls. Minimum inhibitory concentrations were low for all metals (monomethyl mercury, less than 0.05 microgram liter-1; mercuric chloride, less than 1 microgram liter-1; tributyltin chloride, less than 5 micrograms liter-1) except stannic chloride (5 mg liter-1). In some cases, mercuric chloride and monomethyl mercury were equally toxic at comparable concentrations. The Chesapeake Bay community appeared to be slightly more sensitive to metal stress than the Baltimore Harbor community, but this was not true for all treatments or assays. For culturable bacteria the opposite result was found. Thymidine incorporation and glutamate metabolism were much more sensitive indicators of metal toxicity than was viability. To our knowledge, this is the first use of the thymidine incorporation method for ecotoxicology studies. We found it the easiest and fastest of the three methods; it is at least equal in sensitivity to metabolic measurements, and it likely measures the effects on greater portion of the natural community.     

A structural basis for the unequal sensitivity of the major cardiac and liver gap junctions to intracellular acidification: the carboxyl tail length.

The regulation of junctional conductance (Gi) of the major cardiac (connexin43; Cx43) and liver (connexin32; Cx32) gap junction proteins by intracellular hydrogen ion concentration (pH; pHi), as well as well as that of a truncation mutant of Cx43 (M257) with 125 amino acids deleted from the COOH terminus, was characterized in pairs of Xenopus laevis oocytes expressing homologous channels. Oocytes were injected with 40 nl mRNAs (2 micrograms/microliters) encoding the respective proteins; subsequently, cells were stripped, paired, and incubated for 20-24 h. Gj was measured in oocyte pairs using the dual electrode voltage-clamp technique, while pHi was recorded simultaneously in the unstimulated cell by means of a proton-selective microelectrode. Because initial experiments showed that the pH-sensitive microelectrode responded more appropriately to acetate than to CO2 acidification, oocytes expressing Cx32 and wild type and mutant Cx43 were exposed to a sodium acetate saline, which was balanced to various levels of pH using NaOH and HCl. pH was changed in a stepwise manner, and quasi-steady-state Gj -pHi relationships were constructed from data collected at each step after both Gj and pHi had reached their respective asymptotic values. A moderate but significant increase of Gj was observed in Cx43 pairs as pHi decreased from 7.2 to 6.8. In both Cx32 and M257 pairs, Gj increased significantly over a wider pH range (i.e., between 7.2 and 6.3). Further acidification reversibly reduced Gj to zero in all oocyte pairs. Pooled data for the individual connexins obtained during uncoupling were fitted by the Hill equation; apparent 50%-maximum (pK;pKa) values were 6.6 and 6.1 for Cx43 and Cx32, respectively, and Hill coefficients were 4.2 for Cx43 and 6.2 for Cx32. Like Cx32, M257 had a more acidic pKa (6.1) and steeper Hill coefficient (6.0) than wild type Cx43. The pKa and Hill coefficient of M257 were very similar to those of Cx32. These experiments provide the first direct comparison of the effects of acidification on Gj in oocyte pairs expressing Cx43 or Cx32. The results indicate that structural differences in the connexins are the basis for their unequal sensitivity to intracellular acidification in vivo. The data further suggest that a common pH gating mechanism may exist between amino acid residues 1 and 256 in both Cx32 and Cx43. However, the longer carboxyl tail of Cx43 relative to Cx32 or M257 provides additional means to facilitate acidification-induced gating; its presence shifts the pKa from 6.1 (Cx32 and M257) to 6.6 (Cx43) in the conductance of these channels.