Impact of simplifications on numerical modelling of the shallow subsurface at city-scale and implications for shallow geothermal potential.

Anthropogenic infrastructures in the shallow subsurface, such as heated basements, tunnels or shallow geothermal systems, are known to increase ground temperatures, particularly in urban areas. Numerical modelling helps inform on the extent of thermal influence of such structures, and its potential uses. Realistic modelling of the subsurface is often computationally costly and requires large amounts of data which is often not readily available, necessitating the use of modelling simplifications. This work presents a case-study on the city centre of Cardiff, UK, for which high resolution data is available, and compares modelling results when three key modelling components (namely ground elevation, hydraulic gradient distribution and basement geometry) are implemented either 'realistically', i.e. with high resolution data, or 'simplified', utilising commonly accepted modelling assumptions. Results are presented at a point (local) scale and at a domain (aggregate) scale to investigate the impacts such simplifications have on model outputs for different purposes. Comparison to measured data at individual locations shows that the accuracy of temperature outputs from numerical models is largely insensitive to simplification of the hydraulic gradient distribution implemented, while changes in basement geometry affect accuracy of the mean temperature predicted at a point by as much as 3.5 °C. At the domain scale, ground temperatures within the first 20 m show a notable increase (approximately 1 °C volume-averaged and 0.5 °C surface-averaged), while the average heat flux over the domain is about 0.06 W/m2 at 20 m depth. These increased temperatures result in beneficial conditions for shallow geothermal utilisation, producing drilling cost savings of around £1700 per typical household system or about 9% increase in thermal energy potential. Simplifications of basement geometry and (to a lesser degree) the hydraulics can result in an overestimation of these temperatures and therefore over-predict geothermal potential, while the elevation simplification showed little impact.

Metformin in the treatment of non-alcoholic steatohepatitis: a pilot open label trial.

BACKGROUND: Insulin sensitizing agents may be useful in treatment of non-alcoholic fatty liver disease. AIM: A pilot study to evaluate the efficacy and safety of metformin in non-alcoholic fatty liver disease. METHODS: In an open labelled study, patients with histologically confirmed non-alcoholic fatty liver disease were given metformin (20 mg/kg) for 1 year. Insulin resistance (by log homeostasis assessment model analysis for insulin resistance and Quantitative Insulin Sensitivity Check Index) and post-treatment hepatic histology were compared with pre-treatment histology. RESULTS: Fifteen patients completed 1 year of treatment. During the initial 3 months, there was improvement in alanine aminotransferase and aspartate aminotransferase (P-value 0.01 and 0.02, respectively) along with improvement in insulin sensitivity. However, after 3 months, there was no further improvement in insulin sensitivity and there was gradual rise in aspartate aminotransferase and alanine aminotransferase back to pre-treatment levels. Among the 10 patients with post-treatment biopsy, three (33%), showed improvement in steatosis, two (20%) showed improvement in inflammation score and one (10%) showed improvement in fibrosis. CONCLUSION: Metformin treatment was associated with only a transient improvement in liver chemistries. A progressive, sustainable reduction in insulin sensitivity was not noted during treatment.

GroEL/GroES-mediated folding of a protein too large to be encapsulated.

The chaperonin GroEL binds nonnative proteins too large to fit inside the productive GroEL-GroES cis cavity, but whether and how it assists their folding has remained unanswered. We have examined yeast mitochondrial aconitase, an 82 kDa monomeric Fe(4)S(4) cluster-containing enzyme, observed to aggregate in chaperonin-deficient mitochondria. We observed that aconitase folding both in vivo and in vitro requires both GroEL and GroES, and proceeds via multiple rounds of binding and release. Unlike the folding of smaller substrates, however, this mechanism does not involve cis encapsulation but, rather, requires GroES binding to the trans ring to release nonnative substrate, which likely folds in solution. Following the phase of ATP/GroES-dependent refolding, GroEL stably bound apoaconitase, releasing active holoenzyme upon Fe(4)S(4) cofactor formation, independent of ATP and GroES.

A mass on breast imaging predicts coexisting invasive carcinoma in patients with a core biopsy diagnosis of ductal carcinoma in situ.

An image-guided core-needle breast biopsy (IGCNBB) diagnosis of ductal carcinoma in situ (DCIS) is often upgraded to invasive carcinoma (IC) after complete excision. When IC is identified after excision patients must be returned to the operating room for evaluation of their axillary nodes. We performed this study to identify histologic or mammographic features that would predict the presence of invasion when DCIS is documented by IGCNBB. Patients with an IGCNBB diagnosis of DCIS were identified from a prospective database. Imaging abnormalities were classified as either calcification only or mass with or without calcifications. IGCNBB specimens were reviewed to document nuclear grade and the presence of comedo-type necrosis, periductal fibrosis, and periductal inflammation. Patients were divided into two groups, DCIS and IC, on the basis of the final diagnosis after complete excision. From July 1993 through May 2000, 148 of 2995 (4.9%) IGCNBBs demonstrated DCIS; eight were excluded after pathologic review. Of the remaining 140 patients 36 (26%) demonstrated IC after complete excision. The presence of a mass on breast imaging was the only significant predictor of IC (P = 0.04). On the basis of the results of this study we now perform sentinel lymph node mapping and biopsy at the initial surgical procedure for patients with an IGCNBB diagnosis of DCIS and an associated mass on breast imaging.

Biopsy technique has no impact on local recurrence after breast-conserving therapy.

Image-guided core needle breast biopsy (IGCNBB) is an incisional biopsy technique that has been associated with tumor cell displacement. Theoretically tumor cell displacement may affect local recurrence rates in patients treated with breast-conserving therapy (BCT). We performed a study to determine if the biopsy method impacted local control rates following BCT. Patients with nonpalpable breast cancer (invasive and intraductal) diagnosed at our institution and treated with BCT between July 1993 and July 1996 were selected to provide a follow-up period in which the majority of local recurrences should be detected. Patients were divided into two groups based on their method of diagnosis. Group I patients were diagnosed by IGCNBB and group II patients were diagnosed by wire localized excisional breast biopsy (WLEBB). Factors potentially affecting local recurrence rates were retrospectively reviewed. Two hundred eleven patients were treated with BCT, 132 were diagnosed by IGCNBB and 79 by WLEBB. The two patient groups were similar when compared for prognostic factors and treatment. All patients' BCT included histologically negative margins. There were 4 (3.0%) local recurrences in Group I at a median follow-up of 44.4 months and 2 (2.5%) local recurrences in group II at a median follow-up of 50.1 months. This difference was not significant. Breast cancer patients diagnosed by IGCNBB can be treated by BCT with acceptable local control rates. Additional surveillance of our institutional experience and others' is mandatory to validate IGCNBB as the preferred biopsy method for nonpalpable mammographic abnormalities.

Low-density lipoprotein receptor-related protein-5 binds to Axin and regulates the canonical Wnt signaling pathway.

To understand how the Wnt coreceptor LRP-5 is involved in transducing the canonical Wnt signals, we identified Axin as a protein that interacts with the intracellular domain of LRP-5. LRP-5, when expressed in fibroblast cells, showed no effect on the canonical Wnt signaling pathway by itself, but acted synergistically with Wnt. In contrast, LRP-5 mutants lacking the extracellular domain functioned as constitutively active forms that bind Axin and that induce LEF-1 activation by destabilizing Axin and stabilizing beta-catenin. Addition of Wnt caused the translocation of Axin to the membrane and enhanced the interaction between Axin and LRP-5. In addition, the LRP-5 sequences involved in interactions with Axin are required for LEF-1 activation. Thus, we conclude that the binding of Axin to LRP-5 is an important part of the Wnt signal transduction pathway.

ATP-bound states of GroEL captured by cryo-electron microscopy.

The chaperonin GroEL drives its protein-folding cycle by cooperatively binding ATP to one of its two rings, priming that ring to become folding-active upon GroES binding, while simultaneously discharging the previous folding chamber from the opposite ring. The GroEL-ATP structure, determined by cryo-EM and atomic structure fitting, shows that the intermediate domains rotate downward, switching their intersubunit salt bridge contacts from substrate binding to ATP binding domains. These observations, together with the effects of ATP binding to a GroEL-GroES-ADP complex, suggest structural models for the ATP-induced reduction in affinity for polypeptide and for cooperativity. The model for cooperativity, based on switching of intersubunit salt bridge interactions around the GroEL ring, may provide general insight into cooperativity in other ring complexes and molecular machines.

Human Kaposi's sarcoma cell-mediated tumorigenesis in human immunodeficiency type 1 tat-expressing transgenic mice.

BACKGROUND: The human immunodeficiency virus type 1 (HIV-1) transactivator (Tat) protein has been linked to the development and course of Kaposi's sarcoma (KS) associated with acquired immunodeficiency disease syndrome (AIDS-KS). Tat is an 86-101 amino-acid protein encoded by two exons. To evaluate the growth-promoting effects of Tat in AIDS-KS in vivo, we developed transgenic mice expressing the one-exon-encoded 72 amino-acid protein (Tat(72)) and the two-exon-encoded 86 amino-acid protein (Tat(86)). METHODS: Human KS SLK cells were injected subcutaneously into CD4(+) T-cell-depleted male mice, and the tumors that formed after 3-4 weeks were recovered and analyzed for the expression of Tat protein(s), different cytokine messenger RNAs (mRNAs), and matrix metalloproteinases (MMPs). All statistical tests were two-sided. RESULTS: The average tumor weight was maximum in Tat(86) mice ( approximately 600 mg) compared with Tat(72) ( approximately 200 mg) and nontransgenic ( approximately 100 mg) mice (P<.005). Histologic examination of tumors showed spindle-shaped SLK cells with prominent infiltrates of inflammatory cells. All of the tumors from Tat mice expressed abundant Tat mRNA, suggesting that the infiltrating mouse cells actively expressed Tat. A comparison of the growth-promoting cytokines in the tumors from Tat(86)-transgenic and nontransgenic mice showed that the expression of the following cytokines was substantially increased in the tumors of the Tat(86) mice: tumor necrosis factor-alpha, interleukin 6, interleukin 8, granulocyte-macrophage colony-stimulating factor, and basic fibroblast growth factor. Furthermore, these tumors showed abundant expression of a 105-kd MMP activity associated with infiltrates of host leukocytes in the lesions. CONCLUSION: Our in vivo data clearly suggest that extracellular Tat can contribute to the growth and tumorigenesis of human KS cells.

Evidence that beta-tubulin induces a conformation change in the cytosolic chaperonin which stabilizes binding: implications for the mechanism of action.

The class II chaperonin CCT facilitates protein folding by a process that is not well-understood. One striking feature of this chaperonin is its apparent selectivity in vivo, folding only actin, tubulin, and several other proteins. In contrast, the class I chaperonin GroEL is thought to facilitate the folding of many proteins within Escherichia coli. It has been proposed that this apparent selectivity is associated with certain regions of a substrate protein's primary structure. Using limiting amounts of beta-tubulin, beta-tubulin mutants, and beta-tubulin/ftsZ chimeras, we assessed the contribution of select regions of beta-tubulin to CCT binding. In a complementary study, we investigated inter-ring communication in CCT where we exploited polypeptide binding sensitivity to nucleotide to quantitate nucleotide binding. beta-Tubulin bound with a high apparent affinity to CCT in the absence of nucleotide (apparent K(D) approximately 3 nM; its apparent binding free energy, DeltaG, ca. -11.8 kcal/mol). Despite this, the interactions appear to be weak and distributed throughout much of the sequence, although certain sites ("hot spots") may interact somewhat more strongly with CCT. Globally averaged over the beta-tubulin sequence, these interactions appear to contribute ca. -9 to -11 cal/mol per residue, and to account for no more than 50-60% of the total binding free energy. We propose that a conformation change or deformation induced in CCT by substrate binding provides the missing free energy which stabilizes the binary complex. We suggest that by coupling CCT deformation with polypeptide binding, CCT avoids the need for high "intrinsic" affinities for its substrates. This strategy allows for dynamic interactions between chaperonin and bound substrate, which may facilitate folding on the interior surface of CCT in the absence of nucleotide and/or productive release of bound polypeptide into the central cavity upon subsequent MgATP binding. CCT displayed negative inter-ring cooperativity like GroEL. When ring 1 of CCT bound MgATP or beta-tubulin, the affinity of ring 2 for polypeptide or nucleotide was apparently reduced approximately 100-fold.

Multivalent binding of nonnative substrate proteins by the chaperonin GroEL.

The chaperonin GroEL binds nonnative substrate protein in the central cavity of an open ring through exposed hydrophobic residues at the inside aspect of the apical domains and then mediates productive folding upon binding ATP and the cochaperonin GroES. Whether nonnative proteins bind to more than one of the seven apical domains of a GroEL ring is unknown. We have addressed this using rings with various combinations of wild-type and binding-defective mutant apical domains, enabled by their production as single polypeptides. A wild-type extent of binary complex formation with two stringent substrate proteins, malate dehydrogenase or Rubisco, required a minimum of three consecutive binding-proficient apical domains. Rhodanese, a less-stringent substrate, required only two wild-type domains and was insensitive to their arrangement. As a physical correlate, multivalent binding of Rubisco was directly observed in an oxidative cross-linking experiment.

Interaction among GSK-3, GBP, axin, and APC in Xenopus axis specification.

Glycogen synthase kinase 3 (GSK-3) is a constitutively active kinase that negatively regulates its substrates, one of which is beta-catenin, a downstream effector of the Wnt signaling pathway that is required for dorsal-ventral axis specification in the Xenopus embryo. GSK-3 activity is regulated through the opposing activities of multiple proteins. Axin, GSK-3, and beta-catenin form a complex that promotes the GSK-3-mediated phosphorylation and subsequent degradation of beta-catenin. Adenomatous polyposis coli (APC) joins the complex and downregulates beta-catenin in mammalian cells, but its role in Xenopus is less clear. In contrast, GBP, which is required for axis formation in Xenopus, binds and inhibits GSK-3. We show here that GSK-3 binding protein (GBP) inhibits GSK-3, in part, by preventing Axin from binding GSK-3. Similarly, we present evidence that a dominant-negative GSK-3 mutant, which causes the same effects as GBP, keeps endogenous GSK-3 from binding to Axin. We show that GBP also functions by preventing the GSK-3-mediated phosphorylation of a protein substrate without eliminating its catalytic activity. Finally, we show that the previously demonstrated axis-inducing property of overexpressed APC is attributable to its ability to stabilize cytoplasmic beta-catenin levels, demonstrating that APC is impinging upon the canonical Wnt pathway in this model system. These results contribute to our growing understanding of how GSK-3 regulation in the early embryo leads to regional differences in beta-catenin levels and establishment of the dorsal axis.

Benign diagnosis by image-guided core-needle breast biopsy.

Image-guided core-needle breast biopsy (IGCNBB) is widely used to evaluate patients with abnormal mammograms; however, information is limited regarding the reliability of a benign diagnosis. The goal of this study was to demonstrate that a benign diagnosis obtained by IGCNBB is accurate and amenable to mammographic surveillance. Records of all patients evaluated by IGCNBB from July 1993 through July 1996 were reviewed. Biopsies were classified as malignant, atypical, or benign. All benign cases were followed by surveillance mammography beginning 6 months after IGCNBB. Of the 1110 patients evaluated by IGCNBB during the study period, 855 revealed benign pathology. A total of 728 of the 855 patients (85%) complied with the recommendation for surveillance mammography. A total of 196 IGCNBBs were classified as malignant; 59 cases were classified as atypical. The atypical cases were excluded from the statistical analysis. Only two patients have demonstrated carcinoma after a benign IGCNBB during the 2-year minimum follow-up period. The sensitivity and specificity of a benign result were 100.0 and 98.9 per cent, respectively. A benign diagnosis obtained by IGCNBB is accurate and therefore amenable to mammographic surveillance. The results of this study support IGCNBB as the preferred method of evaluating women with abnormal mammograms.

Lessons learned from the initial 100 patient experience with sentinel lymph node mapping in the evaluation of breast cancer.

The initial reports of sentinel lymph node mapping for breast cancer currently appearing in the surgical literature are demonstrating the practicality and accuracy of the technique to evaluate patients for axillary nodal disease. We reviewed our initial 100 patient experience with sentinel node mapping to evaluate our ability to employ this technique in breast cancer patients. We combined a peritumoral injection of a radioactive substance and blue dye. Each sentinel node was evaluated with frozen section analysis, hematoxylin and eosin staining, and, if still negative, five re-cuts were taken from deeper levels of the node and evaluated for immunohistochemical evidence of cytokeratin staining. Sentinel node(s) were identified in all but two patients with 51% demonstrating metastasis. We have demonstrated the ability to accurately perform sentinel node mapping in the evaluation of our breast cancer patients. This exciting advance should become a standard part of breast cancer surgery.

Recurrent juvenile-onset laryngotracheal papillomatosis with transformation to squamous cell carcinoma of the lung.

A 47-year-old female patient with recurrent juvenile-onset laryngotracheal papillomatosis for 27 years had multiple bilateral pulmonary lesions, the largest of which was a well-differentiated squamous cell carcinoma. This case is unique because the malignant transformation occurred in a nonirradiated, nonsmoking patient.

Protection against 3'-to-5' RNA decay in Bacillus subtilis.

A 320-nucleotide RNA with several characteristic features was expressed in Bacillus subtilis to study RNA processing. The RNA consisted of a 5'-proximal sequence from bacteriophage SP82 containing strong secondary structure, a Bs-RNase III cleavage site, and the 3'-proximal end of the ermC transcriptional unit. Comparison of RNA processing in a wild-type strain and a strain in which the pnpA gene, coding for polynucleotide phosphorylase (PNPase), was deleted, as well as in vitro assays of phosphate-dependent degradation, showed that PNPase activity could be stalled in vivo and in vitro. Analysis of mutations in the SP82 moiety mapped the block to PNPase processivity to a particular stem-loop structure. This structure did not provide a block to processivity in the pnpA strain, suggesting that it was specific for PNPase. An abundant RNA with a 3' end located in the ermC coding sequence was detected in the pnpA strain but not in the wild type, indicating that this block is specific for a different 3'-to-5' exonuclease. The finding of impediments to 3'-to-5' degradation, with specificities for different exonucleases, suggests the existence of discrete intermediates in the mRNA decay pathway.

Radial scar of the breast: radiologic-pathologic correlation in 22 cases.

Twenty-two cases were reviewed in which the diagnosis of radial scar (complex sclerosing lesion) of the breast was suspected preoperatively. At mammography, the lesions had a "black star" appearance with long, thin spicules radiating from a radiolucent central area. Excisional rather than core needle biopsy was recommended in all cases. In 13 of 22 cases, including one case of atypical ductal hyperplasia, the lesions proved benign at pathologic analysis. The remaining nine cases were malignant and included one case with a low-nuclear-grade cribriform and micropapillary ductal carcinoma in situ adjacent to the lesion. Results of this study confirm the previously reported association of atypical ductal hyperplasia and carcinoma with radial scar. Furthermore, they demonstrate that a mammographic finding suggestive of radial scar may represent a malignancy that mimics the typical imaging findings in these entities. In cases of mammographically suspected radial scar, all members of the management team as well as the patient should be made aware preoperatively of the potential for benign as well as malignant pathologic findings.

Microsatellite instability and 8p allelic imbalance in stage B2 and C colorectal cancers.

BACKGROUND: Microsatellite instability (MSI) and allelic imbalance involving chromosome arms 5q, 8p, 17p, and 18q are genetic alterations commonly found in colorectal cancer. We investigated whether the presence or absence of these genetic alterations would allow stratification of patients with Astler-Coller stage B2 or C colorectal cancer into favorable and unfavorable prognostic groups. METHODS: Tumors from 508 patients were evaluated for MSI and allelic imbalance by use of 11 microsatellite markers located on chromosome arms 5q, 8p, 15q, 17p, and 18q. Genetic alterations involving each of these markers were examined for associations with survival and disease recurrence. All P values are two-sided. RESULTS: In univariate analyses, high MSI (MSI-H), i.e., MSI at 30% or more of the loci examined, was associated with improved survival (P =.02) and time to recurrence (P =.01). The group of patients whose tumors exhibited allelic imbalance at chromosome 8p had decreased survival (P =.02) and time to recurrence (P =.004). No statistically significant associations with survival or time to recurrence were observed for markers on chromosome arms 5q, 15q, 17p, or 18q. In multivariate analyses, MSI-H was an independent predictor of improved survival (hazard ratio [HR] = 0.51; 95% confidence interval [CI] = 0.31-0.82; P =.006) and time to recurrence (HR = 0.42; 95% CI = 0.24-0.74; P =.003), and 8p allelic imbalance was an independent predictor of decreased survival (HR = 1.89; 95% CI = 1.25-2.83; P =. 002) and time to recurrence (HR = 2.07; 95% CI = 1.32-3.25; P =.002). CONCLUSIONS: Patients whose tumors exhibited MSI-H had a favorable prognosis, whereas those with 8p allelic imbalance had a poor prognosis; both alterations served as independent prognostic factors. To our knowledge, this is the first report of an association between 8p allelic imbalance and survival in patients with colorectal cancer.

Discontinuous appendiceal involvement in ulcerative colitis: pathology and clinical correlation.

Continuous mucosal involvement from the rectum proximally is one of the hallmarks of ulcerative colitis. However, recent pathologic series report appendiceal ulcerative colitis in the presence of a histologically normal cecum, representing a "skip" lesion. The clinical significance of this finding has not been established. Eighty patients, 54 males and 26 females, average age 37.9 years (range 14 to 82 years) who underwent proctocolectomy for ulcerative colitis from January 1990 to September 1995 were examined to determine the rate of discontinuous appendiceal involvement. Excluded were 12 patients with prior appendectomy and 11 with fibrotic obliteration of the appendiceal lumen. Of the remaining 57 patients, seven (12.3%) had clear appendiceal involvement in the presence of a histologically normal cecum. These seven patients clinically were indistinguishable from the 50 patients without skip involvement of the appendix in terms of age at surgery, pretreatment medications, type of surgery, interval from diagnosis to definitive procedure, complications, functional results, and clinical course. Discontinuous appendiceal involvement was found in 12.3% of patients undergoing proctocolectomy for ulcerative colitis, and clinically these patients behave as those without this feature.

Axin and Frat1 interact with dvl and GSK, bridging Dvl to GSK in Wnt-mediated regulation of LEF-1.

Wnt proteins transduce their signals through dishevelled (Dvl) proteins to inhibit glycogen synthase kinase 3beta (GSK), leading to the accumulation of cytosolic beta-catenin and activation of TCF/LEF-1 transcription factors. To understand the mechanism by which Dvl acts through GSK to regulate LEF-1, we investigated the roles of Axin and Frat1 in Wnt-mediated activation of LEF-1 in mammalian cells. We found that Dvl interacts with Axin and with Frat1, both of which interact with GSK. Similarly, the Frat1 homolog GBP binds Xenopus Dishevelled in an interaction that requires GSK. We also found that Dvl, Axin and GSK can form a ternary complex bridged by Axin, and that Frat1 can be recruited into this complex probably by Dvl. The observation that the Dvl-binding domain of either Frat1 or Axin was able to inhibit Wnt-1-induced LEF-1 activation suggests that the interactions between Dvl and Axin and between Dvl and Frat may be important for this signaling pathway. Furthermore, Wnt-1 appeared to promote the disintegration of the Frat1-Dvl-GSK-Axin complex, resulting in the dissociation of GSK from Axin. Thus, formation of the quaternary complex may be an important step in Wnt signaling, by which Dvl recruits Frat1, leading to Frat1-mediated dissociation of GSK from Axin.