An evaluation of injury photography practices in Sexual Assault Referral Centres (SARC).

A review was carried out of injury photography methods in a Sexual Assault Referral Centre (SARC). An initial case study review was conducted with a London based SARC where injury photography of sexual assault complainants was routinely undertaken, and has been for over six years. These images are taken by medical staff using 'point and shoot' compact cameras. Following this and to gain an impression of the national picture, a short survey was sent to 58 SARCs to determine the use of photography in capturing injuries, noting the regularity of its use and the specific types of photographic equipment used. 40 out of the 58 SARCS replied and responses show only three routinely use digital photography as a means to document bodily images. The technique was employed, but not routinely or only infrequently, at a further 20 SARCs. In a further seven centres digital photography was only used for self-referral cases to document general body images, the client's being photographed by the police photographers in any case with police involvement. Five SARCs organised training for their examiners on a Digital Single Lens Reflex (DSLR) photography course, none of which subsequently use their DSLR camera to document bodily injuries. Interestingly, two SARCs report using a colposcope as their camera when they consider that photography 'can't wait' as, for example, when they consider the injury may well disappear by the time police photography can be arranged. The review suggests compact cameras require shorter training, are cheaper to buy, easier to use and produce images suitable for use in court.

Replacing natural wetlands with stormwater management facilities: Biophysical and perceived social values.

Urban expansion replaces wetlands of natural origin with artificial stormwater management facilities. The literature suggests that efforts to mimic natural wetlands in the design of stormwater facilities can expand the provision of ecosystem services. Policy developments seek to capitalize on these improvements, encouraging developers to build stormwater wetlands in place of stormwater ponds; however, few have compared the biophysical values and social perceptions of these created wetlands to those of the natural wetlands they are replacing. We compared four types of wetlands: natural references sites, natural wetlands impacted by agriculture, created stormwater wetlands, and created stormwater ponds. We anticipated that they would exhibit a gradient in biodiversity, ecological integrity, chemical and hydrologic stress. We further anticipated that perceived values would mirror measured biophysical values. We found higher biophysical values associated with wetlands of natural origin (both reference and agriculturally impacted). The biophysical values of stormwater wetlands and stormwater ponds were lower and indistinguishable from one another. The perceived wetland values assessed by the public differed from the observed biophysical values. This has important policy implications, as the public are not likely to perceive the loss of values associated with the replacement of natural wetlands with created stormwater management facilities. We conclude that 1) agriculturally impacted wetlands provide biophysical values equivalent to those of natural wetlands, meaning that land use alone is not a great predictor of wetland value; 2) stormwater wetlands are not a substantive improvement over stormwater ponds, relative to wetlands of natural origin; 3) stormwater wetlands are poor mimics of natural wetlands, likely due to fundamental distinctions in terms of basin morphology, temporal variation in hydrology, ground water connectivity, and landscape position; 4) these drivers are relatively fixed, thus, once constructed, it may not be possible to modify them to improve provision of biophysical values; 5) these fixed drivers are not well perceived by the public and thus public perception may not capture the true value of natural wetlands, including those impacted by agriculture.

Glasshouse vs field experiments: do they yield ecologically similar results for assessing N impacts on peat mosses?

• Peat bogs have accumulated more atmospheric carbon (C) than any other terrestrial ecosystem today. Most of this C is associated with peat moss (Sphagnum) litter. Atmospheric nitrogen (N) deposition can decrease Sphagnum production, compromising the C sequestration capacity of peat bogs. The mechanisms underlying the reduced production are uncertain, necessitating multifactorial experiments. • We investigated whether glasshouse experiments are reliable proxies for field experiments for assessing interactions between N deposition and environment as controls on Sphagnum N concentration and production. We performed a meta-analysis over 115 glasshouse experiments and 107 field experiments. • We found that glasshouse and field experiments gave similar qualitative and quantitative estimates of changes in Sphagnum N concentration in response to N application. However, glasshouse-based estimates of changes in production--even qualitative assessments-- diverged from field experiments owing to a stronger N effect on production response in absence of vascular plants in the glasshouse, and a weaker N effect on production response in presence of vascular plants compared to field experiments. • Thus, although we need glasshouse experiments to study how interacting environmental factors affect the response of Sphagnum to increased N deposition, we need field experiments to properly quantify these effects.

Climatic modifiers of the response to nitrogen deposition in peat-forming Sphagnum mosses: a meta-analysis.

Peatlands in the northern hemisphere have accumulated more atmospheric carbon (C) during the Holocene than any other terrestrial ecosystem, making peatlands long-term C sinks of global importance. Projected increases in nitrogen (N) deposition and temperature make future accumulation rates uncertain. Here, we assessed the impact of N deposition on peatland C sequestration potential by investigating the effects of experimental N addition on Sphagnum moss. We employed meta-regressions to the results of 107 field experiments, accounting for sampling dependence in the data. We found that high N loading (comprising N application rate, experiment duration, background N deposition) depressed Sphagnum production relative to untreated controls. The interactive effects of presence of competitive vascular plants and high tissue N concentrations indicated intensified biotic interactions and altered nutrient stochiometry as mechanisms underlying the detrimental N effects. Importantly, a higher summer temperature (mean for July) and increased annual precipitation intensified the negative effects of N. The temperature effect was comparable to an experimental application of almost 4 g N m(-2) yr(-1) for each 1°C increase. Our results indicate that current rates of N deposition in a warmer environment will strongly inhibit C sequestration by Sphagnum-dominated vegetation.

Which common clinical conditions should medical students be able to manage by graduation? A perspective from Australian interns.

The objectives of the study were to report the development of a core curriculum that details the clinical conditions medical students should be able to manage upon graduation; and to canvass the opinion of interns (first-year postgraduate doctors) regarding their perceptions of the level of skill required to manage each condition. Literature relating to core curriculum development and training of junior medical officers was reviewed and stakeholders in the education and training of medical students and junior doctors in the state of New South Wales, Australia (intern supervisors, academics, registrars, nurses and interns) were consulted. The final curriculum spanned 106 conditions, 77 'differentiated' and 29 'undifferentiated'. Four levels of skill at which conditions should potentially be managed were also identified: 'Theoretical knowledge only'; 'Recognize symptoms and signs without supervision'; Initiate preliminary investigations, management and/or treatment without supervision'; and 'Total investigation, management and/or treatment without supervision'. The list of conditions in the curriculum was converted to a survey format and a one-in-two random sample of interns (n = 193) practising in New South Wales who graduated from the state's three medical schools were surveyed regarding the level of skill required for managing each clinical condition at graduation. A total of 51.3% of interns responded to the survey. Interns felt they should be able to initiate preliminary investigation, management and/or treatment for most conditions in the curriculum, with more than half acknowledging this level of management for 53 of the differentiated and 28 of the undifferentiated conditions. It is concluded that developing core curricula in medical education can involve multiple stakeholders, including junior doctors as the consumers of educational experiences. The data gathered may be useful to medical schools revising their curricula.

Molecular analysis of the locus responsible for production of plantaricin S, a two-peptide bacteriocin produced by Lactobacillus plantarum LPCO10.

A 4.5-kb region of chromosomal DNA carrying the locus responsible for the production of plantaricin S, a two-peptide bacteriocin produced by Lactobacillus plantarum LPCO10 (R. Jiménez-Díaz, J. L. Ruiz-Barba, D. P. Cathcart, H. Holo, I. F. Nes, K. H. Sletten, and P. J. Warner, Appl. Environ. Microbiol. 61:4459-4463, 1995), has been cloned, and the nucleotide sequence has been elucidated. Two genes, designated plsA and plsB and encoding peptides alpha and beta, respectively, of plantaricin S, plus an open reading frame (ORF), ORF2, were found to be organized in an operon. Northern blot analysis showed that these genes are cotranscribed, giving a ca. 0.7-kb mRNA, whose transcription start point was determined by primer extension. Nucleotide sequences of plsA and plsB revealed that both genes are translated as bacteriocin precursors which include N-terminal leader sequences of the double-glycine type. The role of ORF2 is unknown at the moment, although it might be expected to encode an immunity protein of the type described for other bacteriocin operons. In addition, several other potential ORFs have been found, including some which may be responsible for the regulation of bacteriocin production. Two of them, ORF8 and ORF14, show strong homology with histidine protein kinase and response regulator genes, respectively, which have been found to be involved in the regulation of the production of other bacteriocins from lactic acid bacteria. A third ORF, ORF5, shows homology with gene agrB from Staphylococcus aureus, which is involved in the mechanism of regulation of the virulence phenotype in this species. Thus, an agr-like regulatory system for the production of plantaricin S is postulated.

Inhibition of IFN-stimulated gene expression and IFN induction of cytolytic resistance to natural killer cell lysis correlate with E1A-p300 binding.

Treatment of target cells with IFN induces resistance to NK cell lysis. This process is blocked by expression of E1A gene products in adenovirus (Ad)-infected and Ad-transformed cells. We compared the ability of adenovirus serotype 5 (Ad5) E1A exon 1 mutants to inhibit the induction of cytolytic resistance by IFN and block IFN-stimulated gene expression with their capacity to bind the cellular proteins p105 (retinoblastoma gene product), p107, and p300. E1A mutants that did not express conserved region 3 (CR3; residues 138-184) or contained deletions in the nonconserved regions between residues 26-35 or 86-120, bound p105, p107, and p300 and were not impaired in their capacity to block IFN-stimulated gene expression or IFN's induction of cytolytic resistance. E1A mutants with deletions in CR2 (residues 121-138) could not bind p105 or p107, but blocked IFN-stimulated gene expression and IFN's induction of cytolytic resistance. In contrast, mutants in CR1 or the N-terminal nonconserved region (residues 2, 4-25, and 48-60), which define E1A's binding site for p300, were unable to block either IFN-stimulated gene expression or IFN's induction of cytolytic resistance. We conclude that E1A's capacity to block both IFN-stimulated gene expression and IFN's induction of cytolytic resistance appears to be transduced through a pathway that involves E1A-p300 binding. The capacity of E1A to block IFN's induction of cytolytic resistance is probably secondary to E1A's more general ability to inhibit IFN-stimulated gene expression.

Transplanted CG4 cells (an oligodendrocyte progenitor cell line) survive, migrate, and contribute to repair of areas of demyelination in X-irradiated and damaged spinal cord but not in normal spinal cord.

In this study, we have examined the behavior of a lac-Z-transfected O- 2A progenitor cell line, CG4, following transplantation into normal and X-irradiated adult rat spinal cord, and we have also addressed the issue of whether CG4 cells transplanted remotely from ethidium bromide-induced demyelinating lesions in both X-irradiated and nonirradiated spinal cord are able to contribute to their repair. Following transplantation into X-irradiated spinal cord, CG4 cells survive, divide, and migrate extensively. The migration occurs mainly within the parenchymal tissue of the cord without preference for white or gray matter. Moreover, CG4 cells migrating away from their point of introduction are able to enter areas of demyelination and remyelinate the demyelinated axons therein. In contrast, when CG4 cells are transplanted into nonirradiated spinal cord, their survival is limited to areas of damage created by the injection procedure. The CG4 cells do not survive in undamaged, nonirradiated spinal cord. When transplanted remotely from areas of demyelination they are unable to traverse intervening areas of normal white matter, although they may enter lesions if transplanted into their close vicinity. These results have important implications for the development of potential therapeutic strategies for the treatment of multifocal demyelinating disorders that are based on glial cell transplantation.

Transcriptional repression by human adenovirus E1A N terminus/conserved domain 1 polypeptides in vivo and in vitro in the absence of protein synthesis.

The human adenovirus E1A 243R protein (243 residues) transcriptionally represses a set of cellular genes that regulate cellular growth and differentiation. We describe two lines of evidence that E1A repression does not require cellular protein synthesis but instead involves direct interaction with a cellular protein(s). First, E1A 243R protein represses an E1A-repressible promoter in the presence of inhibitors of protein synthesis, as shown by cell microinjection-in situ hybridization. Second, E1A 243R protein strongly represses transcription in vitro from promoters of the E1A-repressible genes, human collagenase, and rat insulin type II. Repression in vitro is promoter-specific, and an E1A polypeptide containing only the N-terminal 80 residues is sufficient for strong repression both in vivo and in vitro. By use of a series of E1A 1-80 deletion proteins, the E1A repression function was found to require two E1A sequence elements, one within the nonconserved E1A N terminus, and the second within a portion of conserved region 1 (40-80). These domains have been reported to possess binding sites for several cellular transcription regulators, including p300, Dr1, YY1, and the TBP subunit of TFIID. The in vitro transcription-repression system described here provides a powerful tool for the further analysis of molecular mechanism and the possible role of these cellular factors.

Differentiation of the O-2A progenitor cell line CG-4 into oligodendrocytes and astrocytes following transplantation into glia-deficient areas of CNS white matter.

The in vitro properties of the CG4 cell line have led to its increasing use as a cell line with which to study the behaviour of the O-2A progenitor cell. In this study we have examined the in vivo behaviour of the CG4 cell line following transplantation into areas of adult rat spinal cord white matter which have been permanently depleted of glial cells by the combination of local X-irradiation and direct injection of 0.1% ethidium bromide. Twenty-one days after transplantation, both myelin-forming oligodendrocytes and glial fibrillary acidic protein-positive astrocytes were identified within the lesion, indicating that the CG4 cell line has bipotential differentiation properties when introduced into a pathological environment consisting of demyelinated axons but devoid of oligodendrocytes or astrocytes. In this respect, the CG4 cell line resembles other glial progenitor cell lines that have been transplanted into similar lesions. In some areas of the lesion, remyelination was observed that was similar in extent to that achieved by growth factor-expanded populations of O-2A progenitor cells. The transplant origin of the cell types within the lesion was confirmed by retroviral incorporation of the lacZ marker gene, the expression of which allowed their identification by histochemistry. In conclusion, the in vivo properties of the CG4 cell line make it a highly suitable line with which to study the behaviour of O-2A progenitors following transplantation into normal and damaged CNS.

Transfection of rat dermal papilla cells with a gene encoding a temperature-sensitive polyomavirus large T antigen generates cell lines retaining a differentiated phenotype.

The dermal papilla is a discrete group of cells at the base of the hair follicle and is implicated in controlling the hair growth cycle. Early passage dermal papilla cells can induce hair growth in vivo, but, upon further culturing, this property is lost. In order to study the events occurring in hair induction, a representative dermal papilla cell line was required. We have transfected passage 1 rat vibrissa dermal papilla cells with a polyomavirus large T gene encoding a temperature-sensitive T antigen, and generated permanent cell lines in which the immortalizing function can be switched off by temperature shift. The cells established without crisis, resembled cells in the starting population, and retained the aggregative properties of early passage dermal papilla cells. Growth studies were performed on the immortalized cell lines, which showed that transferring the cells to the restrictive temperature for the large T gene product resulted in cell senescence or quiescence, and changes in morphology. Implantation of cell pellets into the ears of immunologically compatible rats showed that the immortal cells retained hair-inductive ability. Cytokines are believed to have an important role in the control of hair growth. The pattern of cytokine gene expression in the immortal cell lines was compared with early passage dermal papilla cells and a non-hair-inducing dermal papilla cell line, using reverse transcriptase-polymerase chain reaction. Epidermal growth factor, tumour necrosis factor, and interleukin-1a were detected in the immortalized and non-hair-inducing dermal papilla cell lines, but were absent in passage 2 dermal papilla cells. All other cytokines examined were detected in all the cell types under study. These results demonstrate that the polyomavirus large Ttsa-immortalized dermal papilla cell lines are very similar to passage 2 dermal papilla cells and thus provide a good model for hair growth studies. Cytokine expression profiles indicate that the expression of several cytokines may be implicated in hair induction. Further studies are under way to investigate the relationship between cytokine expression and the hair growth cycle.

Multiple pathways for activation of E2A expression in human KB cells by the 243R E1A protein of adenovirus type 5.

Adenovirus type 5 (Ad5) mutant dl520, which produces only the smaller 243 residue (243R) E1A protein, induced efficient production of the viral E2A 72-kDa DNA binding protein (DBP) in human KB cells, but not in human WI38, 143, or HeLa cells. In transient expression assays, the 243R E1A protein induced transcription from the E2 early promoter in KB but not in HeLa cells; there was no transcription from the E3 promoter in either cell line. In KB cells, truncation of the E2 promoter from -285 to -97 basepairs dramatically reduced transactivation by the 243R E1A product but not by wt E1A, suggesting that the 243R protein acts through factors binding in this region. Multiple deletions in both exon 1 and exon 2 of the 243R E1A protein failed to disrupt its ability to induce DBP expression. The possible redundant pathways for this induction are discussed. The multiplicity of these pathways and the fact that they are all inactivated in the WI38 and 143 lines are surprising.

Induction of apoptosis by adenovirus type 5 E1A in rat cells requires a proliferation block.

Infection with Ad5dl520EIB-, an adenovirus producing only the 243 residue E1A protein and lacking the E1B region, caused apoptosis in normal rat kidney (NRK) cells as judged by the production of nucleosomal DNA fragments. Apoptosis occurred only when the cells were growth-inhibited by cell-cell contacts in confluent cultures or by serum starvation and not when they were actively growing. In uninfected cultures, apoptosis also occurred at confluency, but more slowly than after infection. Studies with E1A deletion mutants of dl520E1B- showed that the regions of the E1A protein essential for induction of apoptosis were those in exon 1 required for binding to the cellular proteins p300 and pRb. Mutants defective at inducing apoptosis were previously found to be defective at inducing baby rat kidney cells to synthesize cellular DNA. In our experiments, cells underwent apoptosis when stimulated by E1A to proliferate under conditions where proliferation was blocked. It is possible that it was the proliferation block opposing the induction of proliferation that led directly to apoptosis. Circumstances leading to induction of apoptosis by c-myc (Evan et al., 1992) are similar and can be interpreted in a similar way.

Functional interactions within adenovirus E1A protein complexes.

The transforming potential of adenovirus E1A oncogene products derives largely from the formation of complexes with cellular proteins, including the p105Rb tumor suppressor and a related p107 species, p130 and p300 proteins, and cyclin A (p60cycA). Extensive quantitative analyses using E1A deletion mutants identified unique binding patterns for each of these polypeptides within the amino terminus and conserved regions 1 and 2 (CR1 and CR2) of E1A proteins. A novel protein, termed p400, was found by peptide mapping to be related to p300, and, like p300, to require the E1A amino terminus and a portion of CR1 for binding. p130 was shown to be related to p107, and like p107, to associate with p60cycA. p107, p130 and p105Rb all interacted primarily with CR2, however, sequences within CR1 and the amino terminus were capable of weak interactions and appeared to function cooperatively with CR2 to bind these proteins. Protein kinase activity present in E1A complexes probably derives at least in part from p60cycA-linked p33cdk2 associated with p107 and p130. In vitro phosphorylation of complexes purified by immunoprecipitation resulted in labeling of several proteins. p60cycA was phosphorylated to about the same extent in cyclin A complexes prepared from either AD5- or mock-infected KB cells, however, that of p130 and p107 was dramatically higher in p60cycA complexes from infected cells. p300 was also phosphorylated in complexes prepared using E1A-specific antibodies. Thus one role of E1A proteins in signal transduction and regulation of the cell cycle may be to control the biological activity of p107, p130 and p300 by enhancing their phosphorylation through complex formation.

Adenovirus e1a proteins and transformation (review).

The E1A gene region of human adenovirus can cooperate with a second gene such as adenovirus E1B or activated ras to transform rodent cells oncogenically. On its own, E1A induces quiescent cells to divide, represses cellular differentiation, induces programmed cell death and affects gene expression. E1A acts through its products, two similar nuclear proteins of 289 and 243 residues, which bind to cellular proteins, particularly p300 and a group of interrelated proteins, p130, p107 and pRb. This review describes what is known of the ways E1A interferes with growth regulation by these and other cellular proteins, such as cyclins and transcription factors, so as to bring about oncogenic transformation.

Induction of gene expression by exon 2 of the major E1A proteins of adenovirus type 5.

We have constructed an adenovirus type 5 (Ad5) E1A mutant, dl1119/520, that produces essentially only exon 2 of the major E1A proteins. In infected primary baby rat kidney cells, this mutant induced expression of the E1B 55-kDa protein, and in infected human KB cells, it induced expression of this protein, the E2A 72-kDa protein, and hexon. In KB cells, this mutant grew substantially better than Ad5 dl312, which lacks E1A, and as well as Ad5 dl520, an E1A mutant producing only the 243-residue protein. These results suggest that exon 2 of E1A proteins on its own was able to activate gene expression. We also constructed mutants of dl1119/520, containing small deletions in regions of exon 2 that others found to be associated with effects on the properties of E1A transformants. None of these deletions destroyed gene activation completely, indicating that there may be some redundancy among sequences in exon 2 for inducing gene expression. The two deletions that decreased induction the most, residues 224 to 238 and 255 to 270, were in regions reported to be associated with the expression of a metalloprotease and with enhanced transformation, suggesting that exon 2 may regulate expression of genes governing cell growth. It is remarkable that all sections of E1A proteins, exon 1, the unique region, and exon 2, have now been found to affect gene expression.

Multiple pathways for gene activation in rodent cells by the smaller adenovirus 5 E1A protein and their relevance to growth and transformation.

By immunoprecipitating protein products from virus-infected baby rat kidney (BRK) cells with specific antibodies, we found that the smaller, 243 residue (243R) E1A protein of human adenovirus 5 (Ad5) activated expression of the virus genes for E1B 55K, E2A 72K, E3 19K, hexon, fibre and penton base and the cellular gene for PCNA. The 243R protein also activated the E2A 72K gene in several rodent cell lines. In transient expression assays, this protein trans-activated the E2 early and major late promoters, suggesting that its effect was at least partially transcriptional. Similar assays with mutants of the E2 early promoter suggested that the ATF- and distal E2F-binding sites were required for this activation. Using mutant viruses with deletions in E1A, we found evidence for three separate pathways by which the 243R protein activated gene expression: one depended on sequences in exon 1 required for this protein to bind to p300, a second depended on sequences in exon 1 required for the protein to bind to pRb and the third appeared to be independent of exon 1 altogether and to depend on exon 2. The relative importance of these pathways for activation varied with the gene and cell. We conclude that a major role of E1A in the transformation of BRK cells by Ad5 is to activate specific genes by at least the first two pathways.

Induction of the cell cycle in baby rat kidney cells by adenovirus type 5 E1A in the absence of E1B and a possible influence of p53.

From previous studies on the induction of DNA synthesis in quiescent primary baby rat kidney cells by adenovirus type 5 (Ad5) E1A deletion mutants, we concluded that induction is prevented only when cellular proteins p300 and pRb are both uncomplexed with E1A (J.A. Howe, J.S. Mymryk, C. Egan, P.E. Branton, and S.T. Bayley, Proc. Natl. Acad. Sci. USA 87:5883-5887, 1990). We have now examined induction by these same mutants in virus lacking the E1B region, so that cellular p53 was no longer complexed to the E1B 55-kDa protein. E1A mutants that fail to bind pRb induced DNA synthesis at a significantly lower level in Ad5 lacking E1B than in Ad5 containing E1B. Apparently, therefore, uncomplexed p53 can partially replace p300 in cooperating with pRb to suppress DNA synthesis in baby rat kidney cells.

Role of phosphorylation near the amino terminus of adenovirus type 5 early region 1A proteins.

Human adenovirus early region 1A (E1A) proteins act as transcriptional regulators and function in the control of DNA synthesis and cell transformation. Little is known about how these viral products are functionally regulated. E1A proteins of adenovirus serotype 5 (Ad5) are phosphorylated at several serine residues and previous studies had indicated that both Ser-89 and Ser-219 are substrates for one or more of the cdc2 family of cell cycle kinases. A second residue near the amino terminus, Ser-96, may also be a site. Although phosphorylation of Ser-89 causes a major shift in gel mobility, the effect on E1A biological activity is unclear. In the present studies we have shown by mutational analysis that phosphorylation at Ser-89 also regulates phosphorylation at Ser-96, suggesting that the gel mobility shift is the result of multiple phosphorylation events. Phosphorylation at Ser-89 did not seem to affect E1A-mediated repression of the simian virus 40 enhancer or trans-activation of the E3 promoter significantly, but it did appear to have a modest but significant effect on transformation of primary baby rat kidney cells.