New insight into flavivirus maturation from structure/function studies of the yellow fever virus envelope protein complex.

IMPORTANCE: All enveloped viruses enter cells by fusing their envelope with a target cell membrane while avoiding premature fusion with membranes of the producer cell-the latter being particularly important for viruses that bud at internal membranes. Flaviviruses bud in the endoplasmic reticulum, are transported through the TGN to reach the external milieu, and enter other cells via receptor-mediated endocytosis. The trigger for membrane fusion is the acidic environment of early endosomes, which has a similar pH to the TGN of the producer cell. The viral particles therefore become activated to react to mildly acidic pH only after their release into the neutral pH extracellular environment. Our study shows that for yellow fever virus (YFV), the mechanism of activation involves actively knocking out the fusion brake (protein pr) through a localized conformational change of the envelope protein upon exposure to the neutral pH external environment. Our study has important implications for understanding the molecular mechanism of flavivirus fusion activation in general and points to an alternative way of interfering with this process as an antiviral treatment.

The feasibility and impact of implementing a computer-guided consultation to target health inequality in Asthma.

Greater Manchester has a greater prevalence and worse asthma outcomes than the national average. This study aims to evaluate a digital approach to primary care asthma management and in particular the initial impact of implementing Clinical Decision Support System software in the form of a computer-guided consultation (CGC) in the setting of primary care asthma reviews in deprived areas of Greater Manchester. The CGC (LungHealth Ltd) is an intelligent decision support system ensuring accurate guideline-based staging of asthma and assessment of asthma control with the software subsequently prompting guideline-standard management. Patients on asthma registers in Greater Manchester Primary Care Networks were identified and underwent remote review by nursing staff using the CGC linked directly to the GP clinical system. Three-hundred thirty-eight patients (mean age 59 (SD 17) years; 60% Female) were reviewed. The CGC reported the patient's asthma control to be "Good" in 22%, "Partial" in 6% and "Poor" in 72%. ACT scores were significantly higher in those patients exhibiting "Good" and "Partial" control when compared to those with "Poor" control. The number of steroid courses and hospital admissions in the previous 12 months was significantly lower in those patients exhibiting "Good" and "Partial" control when compared to those with "Poor" control. Nineteen percent were found not to have a personalised asthma management plan during CGC review, which was alerted by the CGC and subsequently, all but 3 patients had this created on review completion (McNemar's test; p < 0.001). 5% were found not to have been prescribed regular inhaled steroid therapy resulting in the operator being alerted by the CGC in all cases. Overall, 44% underwent alteration in asthma therapy following the CGC review with 82% of these representing treatment escalation. An end-to-end digital service solution is feasible for Asthma within primary care and the utilisation of a CGC when conducting primary care asthma reviews increases implementation of guideline-level management thus addressing healthcare inequality while enabling identification of "high risk" asthma patients and guiding appropriate therapy escalation and de-escalation.

Impact of different glycaemic treatment targets on pregnancy outcomes in gestational diabetes.

AIM: With no current randomized trials, we explored the impact of tight compared with standard treatment targets on pregnancy outcomes in gestational diabetes mellitus (GDM). METHODS: This cohort study of singleton births ≥ 28 weeks' gestation was conducted at two major Australian maternity services (2009-2013). Standardized maternal, neonatal and birth outcomes were examined using routine healthcare data and compared for women with GDM at Service One (n = 2885) and Service Two (n = 1887). Services applied different treatment targets: Service One (standard targets, reference group) fasting < 5.5 mmol/l, 2-h postprandial < 7.0 mmol/l; Service Two (tight targets) fasting < 5.0 mmol/l, 2-h postprandial < 6.7 mmol/l. Multivariable regression with propensity score adjustment was used to examine associations between targets and outcomes. RESULTS: GDM prevalence and insulin use were 7.9% and 31% at Service One, and 5.7% and 46% at Service Two. There were no differences in primary outcomes: birthweight > 90th centile [adjusted odds ratio (OR) 1.06, 95% confidence interval (CI) 0.87-1.30] and < 10th centile (OR 0.84, 95% CI 0.70-1.01), or secondary outcomes gestational hypertension, pre-eclampsia, shoulder dystocia or a perinatal composite. Service Two with tight targets had increased induction of labour (OR 3.63, 95% CI 3.17-4.16), elective Caesarean section (OR 1.75, 95% CI 1.37-2.23) and Apgar scores < 7 at 5 min (OR 1.54, 95% CI 1.05-2.25), decreased hypoglycaemia (OR 0.76, 95% CI 0.61-0.94]), jaundice (OR 0.47, 95% CI 0.35-0.63) and respiratory distress (OR 0.68, 95% CI 0.47-0.98). CONCLUSIONS: Tight GDM treatment targets were associated with greater insulin use and no difference in primary birthweight outcomes. The service with tight targets had higher obstetric intervention, lower rates of reported hypoglycaemia, jaundice, respiratory distress and lower Apgar scores. High-quality interventional data are required before tight treatment targets can be implemented.

A glycerophospholipid-specific pocket in the RVFV class II fusion protein drives target membrane insertion.

The Rift Valley fever virus (RVFV) is transmitted by infected mosquitoes, causing severe disease in humans and livestock across Africa. We determined the x-ray structure of the RVFV class II fusion protein Gc in its postfusion form and in complex with a glycerophospholipid (GPL) bound in a conserved cavity next to the fusion loop. Site-directed mutagenesis and molecular dynamics simulations further revealed a built-in motif allowing en bloc insertion of the fusion loop into membranes, making few nonpolar side-chain interactions with the aliphatic moiety and multiple polar interactions with lipid head groups upon membrane restructuring. The GPL head-group recognition pocket is conserved in the fusion proteins of other arthropod-borne viruses, such as Zika and chikungunya viruses, which have recently caused major epidemics worldwide.

Transducing properties of a pre-structured α-helical DPT-peptide containing a short canine adenovirus type 2 E4orf4 PP2A1-binding sequence.

BACKGROUND: Induction of the death pathway resulting from the specific interaction of the PP2A1 phosphatase with adenoviral E4orf4 protein is a promising approach for cancer therapy. With the aim of deregulating tumor pathways, and mimicking E4orf4 anti-cancer signal, we have previously proposed the DPT technology concept, based on design of specific PP1/PP2A interacting penetrating peptides. METHODS: Using biochemical, structural and cell survival experiments, we have characterized new DPT-peptides containing short PP2A binding sequences. RESULTS: We identified overlapping sequences, located within the N-terminal domain E4orf423-46 of canine adenoviral E4orf4 protein, that interact with the PP2A-Bα subunit of PP2A1 holoenzyme. We characterized DPT-E4orf44 and TAT-E4orf44, two bi-partite cell penetrating peptides containing the 12 PP2A1 binding residues of the canine type 2 E4orf427-38 sequence, respectively fused to the DPT-sh1 and TAT shuttle sequences. Surprisingly DPT-E4orf44, in contrast to inactive TAT-E4orf44, adopted a well defined α-helical structure and co-precipitated PP2A1 from HeLa cell extracts. DPT-E4orf44 also internalized streptavidin-HRP and inhibited survival of HeLa cells more efficiently than TAT, TAT-E4orf44 or the previously published anti-tumor TAT-derived peptide shepherdin. DPT-E4orf44 also efficiently inhibited the survival of human adherent transformed cells, including wild type and p53 mutated colonic HCT116 cells, without affecting survival of human non-transformed fibroblasts. CONCLUSIONS: We characterized the transducing properties of a new α-helical DPT-E4orf44 peptide containing a short PP2A-interacting sequence from canine Adenoviral E4orf4 protein. GENERAL SIGNIFICANCE: Our results suggest that α-helical structured DPT peptides specifically interacting with PP2A could be a valuable anti-cancer drug design scaffold.

Long-term effects of intermittent androgen suppression therapy on lean and fat mass: a 33-month prospective study.

BACKGROUND: To examine changes to whole body and regional lean mass (LM) and fat mass (FM) over 33 months of intermittent androgen suppression therapy (IAST). METHODS: Phase II cohort study of 72 prostate cancer patients without metastatic bone disease. Patients received flutamide 250 mg tid and leuprolide 22.5 mg three monthly depot for the 9-month initial treatment phase (iTREAT), at which point patients ceased therapy providing PSA <4 ng ml(-1) with continued monitoring for further 2 years (POST). AST was recommenced when PSA exceeded pretreatment level or ≥ 20 ng ml(-1). Body composition was assessed using dual energy X-ray absorptiometry at baseline, completion of treatment phase, and 1 and 2 years post treatment phase (months 21 and 33). RESULTS: LM decreased by 1.3 kg and FM increased by 2.3 kg (P<0.001) following iTREAT. During the POST period, there were no further adverse effects on LM or FM, but also no recovery to pretreatment levels. Patients who failed to recover testosterone by month 33 experienced a significant increase in FM compared with those who recovered eugonadal levels of testosterone (10 nmol ml(-1); P = 0.019). Change in testosterone was moderately correlated to changes in % FM (r = -0.314, P<0.028) and LM (r = 0.300, P<0.036) during POST phase. Waist circumference progressively increased over time and by 2 years, POST had not recovered to baseline levels. CONCLUSIONS: Loss of LM and gain in FM during the 9-month iTREAT was not reversed during 2-year POST, although further deterioration was not observed. Subgroup analysis identified those recovering testosterone showed some body composition improvements. These findings suggest potential benefits of IAST, where testosterone levels are able to recover, to reduce the ongoing adverse effects on body composition, such as the acceleration of sarcopenia and risks associated with metabolic disease.

Some insights about 1,6-diphenyl-1,3,5-hexatriene-lipid supramolecular assemblies by steady-state fluorescence measurements.

1,6-Diphenyl-1,3,5-hexatriene (DPH) is the most widely proposed molecular probe for the post-column fluorescence derivatization of lipids after liquid chromatography separation. This kind of detection consists of a supramolecular combination of DPH and eluted lipids. The detection is optimally performed in a mainly aqueous environment (over 80% v/v) because the weak fluorescence of DPH in water is drastically enhanced upon formation of supramolecular assemblies with lipids. In the present study, and in order to obtain better spectroscopic insights into the nature of these supramolecular assemblies, two different lipids were tested, 1,2,3-tridodecanoylglycerol (LLL) as a model triglyceride (nonpolar lipid) and dimyristoylphosphatidylcholine (DMPC) as a model phosphatidylcholine (charged amphiphilic lipid). Stoichiometry and association constants were determined on the basis of the variation of fluorescence intensity in the presence of various concentrations of lipids. LLL(60)-DPH(2) and DMPC(200)-DPH(2) complexes were identified with association constants as high as K(2) = (5.8 +/- 0.5) x 10(13) M(-2) and (17.3 +/- 2.0) x 10(13) M(-2) for LLL and DMPC, respectively. The fluorescence intensity of DPH in the presence of LLL is greater than in the presence of DMPC. An attempt to characterize the insertion mode of DPH in the lipidic supramolecular assemblies is also made.

Assessment of population structure by single nucleotide polymorphisms (SNPs) in goat breeds.

Single nucleotide polymorphisms (SNPs) may be used in biodiversity studies and commercial tasks like traceability, paternity testing and selection for suitable genotypes. Twenty-seven SNPs were characterized and genotyped on 250 individuals belonging to eight Italian goat breeds. Multilocus genotype data were used to infer population structure and assign individuals to populations. To estimate the number of groups (K) to test in population structure analysis we used likelihood values and variance of the bootstrap samples, deriving optimal K from a drop in the likelihood and a rise in the variance plots against K.

Proteomic identification of M. tuberculosis protein kinase substrates: PknB recruits GarA, a FHA domain-containing protein, through activation loop-mediated interactions.

Genes for functional Ser/Thr protein kinases (STPKs) are ubiquitous in prokaryotic genomes, but little is known about their physiological substrates and their actual involvement in bacterial signal transduction pathways. We report here the identification of GarA (Rv1827), a Forkhead-associated (FHA) domain-containing protein, as a putative physiological substrate of PknB, an essential Ser/Thr protein kinase from Mycobacterium tuberculosis. Using a global proteomic approach, GarA was found to be the best detectable substrate of the PknB catalytic domain in non-denatured whole-cell protein extracts from M. tuberculosis and the saprophyte Mycobacterium smegmatis. Enzymological and binding studies of the recombinant proteins demonstrate that docking interactions between the activation loop of PknB and the C-terminal FHA domain of GarA are required to enable efficient phosphorylation at a single N-terminal threonine residue, Thr22, of the substrate. The predicted amino acid sequence of the garA gene, including both the N-terminal phosphorylation motif and the FHA domain, is strongly conserved in mycobacteria and other related actinomycetes, suggesting a functional role of GarA in putative STPK-mediated signal transduction pathways. The ensuing model of PknB-GarA interactions suggests a substrate recruitment mechanism that might apply to other mycobacterial kinases bearing multiple phosphorylation sites in their activation loops.

Sudden unexplained infant death in 20 regions in Europe: case control study.

BACKGROUND: After striking changes in rates of sudden unexplained infant death (SIDS) around 1990, four large case-control studies were set up to re-examine the epidemiology of this syndrome. The European Concerted Action on SIDS (ECAS) investigation was planned to bring together data from these and new studies to give an overview of risk factors for the syndrome in Europe. METHODS: We undertook case-control studies in 20 regions. Data for more than 60 variables were extracted from anonymised records of 745 SIDS cases and 2411 live controls. Logistic regression was used to calculate odds ratios (ORs) for every factor in isolation, and to construct multivariate models. FINDINGS: Principal risk factors were largely independent. Multivariately significant ORs showed little evidence of intercentre heterogeneity apart from four outliers, which were eliminated. Highly significant risks were associated with prone sleeping (OR 13.1 [95% CI 8.51-20.2]) and with turning from the side to the prone position (45.4 [23.4-87.9]). About 48% of cases were attributable to sleeping in the side or prone position. If the mother smoked, significant risks were associated with bed-sharing, especially during the first weeks of life (at 2 weeks 27.0 [13.3-54.9]). This OR was partly attributable to mother's consumption of alcohol. Mother's alcohol consumption was significant only when baby bed-shared all night (OR increased by 1.66 [1.16-2.38] per drink). For mothers who did not smoke during pregnancy, OR for bed-sharing was very small (at 2 weeks 2.4 [1.2-4.6]) and only significant during the first 8 weeks of life. About 16% of cases were attributable to bed-sharing and roughly 36% to the baby sleeping in a separate room. INTERPRETATION: Avoidable risk factors such as those associated with inappropriate infants' sleeping position, type of bedding used, and sleeping arrangements strongly suggest a basis for further substantial reductions in SIDS incidence rates.

Effects of seed bank disturbance on the fine-scale genetic structure of populations of the rare shrub Grevillea macleayana.

Dispersal in most plants is mediated by the movement of seeds and pollen, which move genes across the landscape differently. Grevillea macleayana is a rare, fire-dependent Australian shrub with large seeds lacking adaptations for dispersal; yet it produces inflorescences adapted to pollination by highly mobile vertebrates (eg birds). Interpreting fine-scale genetic structure in the light of these two processes is confounded by the recent imposition of anthropogenic disturbances with potentially contrasting genetic consequences: (1) the unusual foraging behaviour of exotic honeybees and 2. widespread disturbance of the soil-stored seedbank by road building and quarrying. To test for evidence of fine-scale genetic structure within G. macleayana populations and to test the prediction that such structure might be masked by disturbance of the seed bank, we sampled two sites in undisturbed habitat and compared their genetic structure with two sites that had been strongly affected by road building using a test for spatial autocorrelation of genotypes. High selfing levels inferred from genotypes at all four sites implies that pollen dispersal is limited. Consistent with this, we observed substantial spatial clustering of genes at 10 m or less in the two undisturbed populations and argue that this reflects the predicted effects of both high selfing levels and limited seed dispersal. In contrast, at the two sites disturbed by road building, spatial autocorrelation was weak. This suggests there has been mixing of the seed bank, counteracting the naturally low dispersal and elevated selfing due to honeybees. Pollination between near neighbours with reduced relatedness potentially has fitness consequences for G. macleayana in disturbed sites.

Plateau 'pop-up' in the great 1897 Assam earthquake.

The great Assam earthquake of 12 June 1897 reduced to rubble all masonry buildings within a region of northeastern India roughly the size of England, and was felt over an area exceeding that of the great 1755 Lisbon earthquake. Hitherto it was believed that rupture occurred on a north-dipping Himalayan thrust fault propagating south of Bhutan. But here we show that the northern edge of the Shillong plateau rose violently by at least 11 m during the Assam earthquake, and that this was due to the rupture of a buried reverse fault approximately 110 km in length and dipping steeply away from the Himalaya. The stress drop implied by the rupture geometry and the prodigious fault slip of 18 +/- 7 m explains epicentral accelerations observed to exceed 1g vertically and surface velocities exceeding 3 m s-1 (ref. 1). This quantitative observation of active deformation of a 'pop-up' structure confirms that faults bounding such structures can penetrate the whole crust. Plateau uplift in the past 2-5 million years has caused the Indian plate to contract locally by 4 +/- 2 mm yr-1, reducing seismic risk in Bhutan but increasing the risk in northern Bangladesh.

A new visual identity for the National Health Service.

The following article gives a brief overview of the new visual identity being adopted by the National Health Service in England. It looks at the thinking behind the identity, the identity's component parts and provides sources for obtaining further information on the identity's application. It is compiled from a presentation by Stephanie Hood from the corporate identity team of the NHS Executive communications unit given on 22nd October 1999 at the National Designers in Health Network seminar, Time-out '99, Sheffield. Supporting information was obtained from the NHS Communications website http:¿nww.doh.nhsweb.uk/commsnet.

Probing the role of a conserved M1 proline residue in 5-hydroxytryptamine(3) receptor gating.

A conserved proline residue is found in the first transmembrane domain (M1) of every subunit in the ligand-gated ion channel superfamily. The position of this proline between the N-terminal extracellular agonist binding and the second transmembrane (M2) channel lining domains in the primary sequence suggests its possible involvement in the gating of the receptor. Replacing this proline with alanine, glycine, or leucine in the 5-hydroxytryptamine (5-HT)(3A) homomeric receptors expressed in Xenopus laevis oocytes resulted in the absence of 5-HT-induced whole-cell currents, although there were normal levels of specific surface [(3)H]granisetron ([(3)H]BRL-43694) binding sites. To determine what properties of the conserved proline are critical for the function of the channel, two imino acids and an alpha-hydroxy acid were incorporated at the proline position using the nonsense suppression method. trans-3-Methyl-proline, pipecolic acid, and leucic acid were able to replace the conserved proline to produce active channels with EC(50) values similar to that for the wild-type receptor. These trends are preserved in the heteromeric receptors consisting of 5-HT(3A) and 5-HT(3B) subunits in oocytes. The prominent common feature among these residues and proline is the lack of hydrogen bond donor activity, potentially resulting in a flexible secondary structure in the M1 region. Thus, lack of hydrogen bond donor activity may be a key element in channel gating and may explain the high degree of conservation of this M1 proline.

A prospective management study of slow-release aspirin in the palliation of uteroplacental insufficiency predicted by uterine artery Doppler at 20 weeks.

OBJECTIVE: To investigate the effect of low-dose, slow-release aspirin in reducing the incidence and/or severity of pregnancy complications in women identified as high risk of developing problems associated with uteroplacental insufficiency, namely pre-eclampsia or delivering a small-for-gestational age (SGA) baby. DESIGN: A prospective, randomized management study. One thousand and twenty-two women of mixed parity underwent color flow/pulsed Doppler (CFPD) imaging of the uterine arteries at the time of the 17-23 week (mean 19.9) anomaly scan. Women who were screen positive were randomized to a control or treatment group. The treatment group was given 100-mg slow-release aspirin (Disprin CV) daily and followed up at regular intervals. Women in the routine group received routine antenatal care. Main outcome measures were pre-eclampsia and SGA < 3rd centile. Secondary outcome measures were: SGA < 10th centile, pre-eclampsia requiring delivery before 34 weeks, placental abruption, an Apgar score < 7 at 5 min, admission to neonatal intensive care unit or a pregnancy that resulted in a stillbirth or neonatal death. Odds ratios (OR) with 95% confidence intervals (CI) were calculated for severe and any complications. RESULTS: Two hundred and sixteen women were screen positive according to the defined criteria. One hundred and three women were assigned to the treatment group and 113 to the control group. The difference in the incidence of pre-eclampsia and SGA < 3rd centile between the control and treatment groups did not reach statistical significance. There was a statistically significant reduction in any (OR 0.41 (CI 0.35-0.45), P < 0.01) and severe pregnancy complications (OR 0.43 (CI 0.21-0.84), P < 0.05) in the treatment group compared with the controls. CONCLUSIONS: The administration of slow-release aspirin to women identified as high risk, using color Doppler imaging of the uterine arteries at 20 weeks' gestation, did not significantly alter the incidence of pre-eclampsia or delivery of a SGA baby. It did, however, improve the outcome by reducing the overall incidence of complications associated with uteroplacental insufficiency.

Neutralizing human anti crotoxin scFv isolated from a nonimmunized phage library.

Combinatorial phage display technology offers a new possibility for making human antibodies which could be used in immune therapy. We explored the use of this technology to make human scFvs specific for crotoxin, the main toxic component of the venom of the South-American rattlesnake Crotalus durissus terrificus. Crotoxin, a phospholipase A2 neurotoxin constituted by the association of two subunits, exerts its lethal action by blocking neuromuscular transmission. This is the first report of human anticrotoxin scFvs (scFv 1, scFv 6 and scFv 8) isolated from a naive library of more than 1010 scFv clones with in vivo neutralizing activity. Nevertheless, differences are observed at the level of biological and immunological effects. Only scFv 8 is able to reduce the myotoxicity induced by crotoxin and scFv 1 is capable of altering the in vitro enzymatic activity of this toxin. All three scFvs recognize a region of one subunit located at the junction with the other one. Moreover these scFvs share strong amino acid homologies at the level of either the heavy or the light chain. Taken together, our results suggest that the use of human anticrotoxin scFvs may lead to a new and less aggressive passive immune therapy against poisoning by the venom of Crotalus durissus terrificus.

Protein engineering of cytochrome p450(cam) (CYP101) for the oxidation of polycyclic aromatic hydrocarbons.

Mutations of the active site residues F87 and Y96 greatly enhanced the activity of cytochrome P450(cam) (CYP101) from Pseudomonas putida for the oxidation of the polycyclic aromatic hydrocarbons phenanthrene, fluoranthene, pyrene and benzo[a]pyrene. Wild-type P450(cam) had low (<0.01 min(-1)) activity with these substrates. Phenanthrene was oxidized to 1-, 2-, 3- and 4-phenanthrol, while fluoranthene gave mainly 3-fluoranthol. Pyrene was oxidized to 1-pyrenol and then to 1,6- and 1,8-pyrenequinone, with small amounts of 2-pyrenol also formed with the Y96A mutant. Benzo[a]pyrene gave 3-hydroxybenzo[a]pyrene as the major product. The NADH oxidation rate of the mutants with phenanthrene was as high as 374 min(-1), which was 31% of the camphor oxidation rate by wild-type P450(cam), and with fluoranthene the fastest rate was 144 min(-1). The oxidation of phenanthrene and fluoranthene were highly uncoupled, with highest couplings of 1.3 and 3.1%, respectively. The highest coupling efficiency for pyrene oxidation was a reasonable 23%, but the NADH turnover rate was slow. The product distributions varied significantly between mutants, suggesting that substrate binding orientations can be manipulated by protein engineering, and that genetic variants of P450(cam) may be useful for studying the oxidation of polycyclic aromatic hydrocarbons by P450 enzymes.

Management of vaginal intraepithelial neoplasia: A series of 132 cases with long-term follow-up.

A series of 132 cases of vaginal intraepithelial neoplasia (VAIN) is presented, including nine (6.8%) where early invasive carcinoma of the vagina was found in the course of initial management of the VAIN. The majority of patients (75%) had high-grade VAIN (two or three). Seventy-two (55%) had undergone a prior hysterectomy; 22 for preinvasive disease (CIN), 33 for invasive gynecological cancer, 13 for benign reasons, and in 4 the reason for the hysterectomy and/or the Pap smear history was not known. Twenty-one (16%) had received prior pelvic radiotherapy. VAIN was noted to involve either the vaginal vault (in the post-hysterectomy group) or the upper vagina (in the no hysterectomy group) in more than 80% cases. A variety of treatment modalities was used with varying degrees of success. For high-grade VAIN excisional treatments had an overall (first-line plus subsequent) cure rate of 69% (53/77). The state of the surgical margins did not correlate with the risk of residual disease. CO2 laser ablation was curative in 69% (18/26) of cases and was significantly better than electrocoagulation diathermy which was curative in only 25% (3/12) of cases (P = 0.013). Five-fluorouracil cream was curative in 46% (5/11) of cases, including four patients who had received prior radiotherapy. Radiotherapy was effective in eradicating VAIN in the two cases where it was used as the primary treatment modality. Progression of high-grade VAIN to invasive cancer occurred in eight (8%) cases; after no treatment in two cases, after treatment failure in five cases, and as a late recurrence in one case. For low-grade VAIN an observational approach after biopsy was initially adopted in eight patients and regression occurred in seven (88%) of these patients. Other miscellaneous treatments were also effective in low-grade VAIN. These data provide evidence that high-grade VAIN is a precursor to invasive cancer of the vagina and every attempt should be made to eradicate it. Based on our experience and a review of the literature we have proposed a plan for optimal management of this condition.

"The birthing from within holistic sphere": a conceptual model for childbirth education.

An expanded conceptual model of childbirth education is offered, proposing the benefits of balancing informative teaching processes with creative, experiential, introspective learning processes for parents. The application of these two teaching dimensions to exploring four different perspectives of birth (the mother's, the father's, the baby's, and the culture's) is discussed, along with examples from "Birthing From Within" classes. Implications for current practice and the evolving role of childbirth educator are noted.