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The physical properties of neutrons emitted from neutron-induced fission are fundamental to our understanding of nuclear fission. However, while state-of-the-art fission models still incorporate isotropic fission neutron spectra, it is believed that the preequilibrium prefission component of these spectra is strongly anisotropic. The lack of experimental guidance on this feature has not motivated incorporation of anisotropic neutron spectra in fission models, though any significant anisotropy would impact descriptions of a fissioning system. In the present work, an excess of counts at high energies in the fission neutron spectrum of ^{239}Pu is clearly observed and identified as an excess of the preequilibrium prefission distribution above the postfission neutron spectrum. This excess is separated from the underlying postfission neutron spectrum, and its angular distribution is determined as a function in incident neutron energy and outgoing neutron detection angle. Comparison with neutron scattering models provides the first experimental evidence that the preequilibrium angular distribution is uncorrelated with the fission axis. The results presented here also impact the interpretation of several influential prompt fission neutron spectrum measurements.

BACKGROUND: Sarcopenia is defined as the loss of muscle mass and function with age and is associated with decline in mobility, frailty, falls and mortality. There is considerable interest in understanding the underlying mechanisms. Our aim was to characterise muscle morphology changes associated with sarcopenia among community dwelling older men.METHODS: One hundred and five men aged 68-76 years were recruited to the Hertfordshire Sarcopenia Study (HSS) for detailed characterisation of muscle including measures of muscle mass, strength and function. Muscle tissue was obtained from a biopsy of the vastus lateralis for 99 men and was processed for immunohistochemical studies to determine myofibre distribution and area, capillarisation and satellite cell (SC) density.RESULTS: Six (6 %) men had sarcopenia as defined by the European Working Group on Sarcopenia in Older People (EWGSOP) criteria. These men had lower SC density (1.7 cells/mm(2) vs 3.8 cells/mm(2), p?=?0.06) and lower SC/fibre ratio (0.02 vs 0.06, p?=?0.06) than men without sarcopenia. Although men with sarcopenia tended to have smaller myofibres and lower capillary to fibre ratio, these relationships were not statistically significant.CONCLUSION: We have shown that there may be altered muscle morphology parameters in older men with sarcopenia. These results have the potential to help identify cell and molecular targets for therapeutic intervention. This work now requires extension to larger studies which also include women.

The European Working Time Directive and the New Deal have decreased the number of hours worked by anaesthetic trainees. We implemented the Working Time Directive in May 2004 and evaluated the effect of its implementation on training. During two 6-month periods, one before and one after the change, we determined the number of operating lists undertaken by each Specialist Registrar in Anaesthesia. After implementation of the Working Time Directive, the mean number of lists performed by Specialist Registrars decreased from 24 to 21 lists per registrar per month, a 13% decrease. Exposure to subspecialty lists was the same in both periods, but this was at the expense of general lists and those in remote locations. We conclude that the Working Time Directive has had a measurable impact on the training of paediatric anaesthetists, but that the significance of this change for clinical practice has not yet been measured.

The structural specificity of vitamin D derivatization by PTAD (4-phenyl-1,2,4-triazoline-3,5-dione) was probed using synthetic analogues and ion trap mass spectrometry. EB 1089, a vitamin D(3) analogue which contains a second site for Diels--Alder cycloaddition on its side-chain, allowed the examination of derivatization modes and comparisons of ion fragment structures. The origins of a PTAD-vitamin D(3) ion fragment, commonly used in metabolite characterization and quantitation of vitamin D(3) analogues (m/z 314), were established; ion trap mass spectrometry revealed that the PTAD comprises a portion of this diagnostic fragment, and is not lost by a retro-Diels--Alder step. Furthermore, the unique structure of the EB 1089 side-chain also permits facile determination of its side-chain metabolism. Use of PTAD derivatization and detection of metabolite-specific ion fragments identify hydroxylation at the end of the EB 1089 sidechain. It is believed that the results from these studies provide a clearer understanding of the mass spectrometry of triazolinedione derivatives, not only in the specific case of EB 1089, but also in their application to other vitamin D compounds.

Cell density effects were investigated on tumorous hormonal secretion from 10 pituitary adenomas: 3 somatotrophinomas secreting GH and PRL; 7 gonadotrophinomas, 3 co-secreted both FSH and LH, all 7 secreted LH. Enzymatically dispersed tissue was plated out in 24-well plates at 5 ×105, 105, 5 ×104 and 104 cells/well in serum-free media. Media were collected weekly for 2 weeks. Results: In 3 of 3 somatotrophinomas, GH and PRL secretion was higher (p < 0.05) at both week 1 and 2 from 104 cells/well, but similar at other cell densities. In all 3 gonadotrophinomas, the FSH secretory rate was highest at 5 ×105 cells/well which fell as cell density decreased. Conversely, in 7 of 7 gonadotrophinomas the LH secretory rate was highest at 104 cells/well (p < 0.01) which fell as cell density increased. Conclusion: These data suggest that paracrine factors may modulate tumorous GH, PRL, FSH and LH secretion, and show that FSH and LH secretion vary inversely as cell density increases.

The objective of the study was to examine the effect of Brahman genetics on collagen enzymatic crosslinking gene expression and meat tenderness. Steers were randomly selected to represent a high percentage Brahman genetics (n = 13), Half-Blood genetics (n = 13), Brangus genetics (n = 13), and a high percentage Angus genetics (n = 13). Muscle samples from the Longissimus lumborum muscle were collected at weaning and harvest and reverse transcription quantitative PCR (qPCR) analysis was conducted to measure the mRNA expression of lysyl oxidase (LOX), bone morphogenetic protein 1 (BMP1), and cystatin C (CYS). Steaks from subject animals were collected at harvest, aged for 14 d and subjected to collagen analysis, Warner-Bratzler Shear Force (WBS) and trained sensory panel analysis (tenderness, juiciness, and connective tissue). Data indicated that Half-Blood and Brahman steers had greater (P0.05) WBS values and tended to receive decreased (P0.06) panel tenderness scores than Angus and Brangus steers. Panelists tended to detect more connective tissue in Brahman and Half-Blood steaks when compared to Angus and Brangus steaks (P0.07). Crosslinking gene expression data revealed that at weaning Half-Blood steers had more (P0.05) mRNA expression of CYS and LOX than Angus and Brangus steers. At weaning and harvest, all genetic groups had similar mRNA expression of BMP1 (P0.10). At harvest, Brangus and Angus steers had greater LOX mRNA expression than Brahman cattle (P0.05). Pearson's correlation coefficients indicated that only weaning CYS mRNA expression was correlated to WBS, panel tenderness and connective tissue scores (P0.05). Expression of LOX was only correlated to these measures at harvest, and BMP1 was correlated to these traits at both time periods (P0.05). These results indicate that collagen crosslinking enzyme activity, as indicated by mRNA levels, early in an animal's life may account for some of the variation seen in steak tenderness due to Brahman genetic influence.

The presence of mRNA transcripts for cytokines in normal and neoplastic human breast tissue has been investigated. Using reverse transcriptase-linked polymerase chain reaction (RT-PCR), we have specifically screened for the following cytokines: interleukin (IL)-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, tumour necrosis factor (TNF)-α, TNF-β and interferon (IFN)-γ. No significant differences in expression of IL-1α, IL-1β, IL-4, IL-6, TNF-α or TNF-β were observed between the 2 groups of tissues. However, there was a significant difference in expression of IL-8 transcripts (p = 0.0017) which was higher in the neoplastic population. Transcripts for IL-2, IL-3, IL-5, IL-7 and IFN-γ were not detected in either group. There was no evidence of associations between cytokine expression and tumour histological grade, patient age or lymph node metastases. Correlating tumour types with specific cytokine transcripts revealed high expression of IL-8, and to a lesser extent, IL-8 and TNF-β irrespective of tumour origin. Analysis of primary epithelial and stromal cultures derived from both types of tissue showed that increased levels of IL-8, but not IL-6, were secreted by cells obtained from tumours. Thus, breast tissue of both normal and neoplastic origin expresses a wide range of cytokines. Increased or aberrant expression of cytokines, in particular IL-8, may be involved in the development/progression of breast cancer. Int. J. Cancer 72:937–941, 1997. © 1997 Wiley-Liss, Inc.

1,25-(OH)2D3 and 24,25-(OH)2D3 mediate their effects on chondrocytes through the classic vitamin D receptor (VDR) as well as through rapid membrane-mediated mechanisms which result in both nongenomic and genomic effects. In intact cells, it is difficult to distinguish between genomic responses via the VDR and genomic and nongenomic responses via membrane-mediated pathways. In this study, we used two hybrid analogues of 1,25-(OH)2D3 which have been modified on the A-ring and C,D-ring side chain (1α-(hydroxymethyl)-3β-hydroxy-20-epi-22-oxa-26,27-dihomo vitamin D3 (analogue MCW-YA = 3a) and 1β-(hydroxymethyl)-3α-hydroxy-20-epi-22-oxa-26,27-dihomo vitamin D3 (analogue MCW-YB = 3b) to examine the role of the VDR in response of rat costochondral resting zone (RC) and growth zone (GC) chondrocytes to 1,25-(OH)2D3 and 24,25-(OH)2D3. These hybrid analogues are only 0.1% as effective in binding to the VDR from calf thymus as 1,25-(OH)2D3. Chondrocyte proliferation ([3H]-thymidine incorporation), proteoglycan production ([35S]-sulfate incorporation), and activity of protein kinase C (PKC) were measured after treatment with 1,25-(OH)2D3, 24,25-(OH)2D3, or the analogues. Both analogues inhibited proliferation of both cell types, as did 1,25-(OH)2D3 and 24,25-(OH)2D3. Analogue 3a had no effect on proteoglycan production by GCs but increased that by RCs. Analogue 3b increased proteoglycan production in both GC and RC cultures. Both analogues stimulated PKC in GC cells; however, neither 3a nor 3b had an effect on PKC activity in RC cells. 1,25-(OH)2D3 and 3a decreased PKC in matrix vesicles from GC cultures, whereas plasma membrane PKC activity was increased, with 1,25-(OH)2D3 having a greater effect. 24,25-(OH)2D3 caused a significant decrease in PKC activity in matrix vesicles from RC cultures; 24,25-(OH)2D3, 3a, and 3b increased PKC activity in the plasma membrane fraction, however. Thus, with little or no binding to calf thymus VDR, 3a and 3b can affect cell proliferation, proteoglycan production, and PKC activity. The direct membrane effect is analogue-specific and cell maturation–dependent. By studying analogues with greatly reduced affinity for the VDR, we have provided further evidence for the existence of a membrane receptor(s) involved in mediating nongenomic effects of vitamin D metabolites. J. Cell. Biochem. 66:457–470, 1997. © 1997 Wiley-Liss, Inc.

A method to determine whether dispersed human anterior pituitary adenoma cells proliferate in mixed culture was developed. Fifteen pituitary adenomas were dispersed enzymatically to single cells, following which twelve were double immunostained after eight days. Proliferating cells were identified immunologically following one hour of bromo-deoxyuridine incorporation. Adenoma cells were subsequently identified with an anti-neuron-specific enolase antibody system. A time course of bromo-deoxyuridine labelling was performed on three nonfunctional adenomas over a four day period, with bromo-deoxyuridine being added to cultures at one hour, 24 hours and four days prior to immunostaining. Double immunolabelled cells were unambiguously identified by a dark brown nucleus surrounded by red cytoplasm. Eight out of 12 pituitary adenomas (two prolactinomas, three nonfunctional, three growth hormone secreting) showed an increased bromo-deoxyuridine labelling index (range 0.1%-1.4%). Bromo-deoxyuridine incorporation over four days showed an increase in bromo-deoxyuridine from 0.02%, 0.03% and 3.3% at one hour to 10.1%, 1.3% and 5.0% at four days, respectively, but evidence of mitosis was scant. This study shows that pituitary adenomas may proliferate in vitro and that this double immunostaining method may be used as an in vitro proliferation assay in a mixed cell population.

✓ The authors compared detection methods for cell proliferation in human anterior pituitary adenomas using histological sections and dispersed cell culture. After tumor cells had been grown for 4 days in dispersed culture, bromodeoxyuridine (BUdR), proliferating cell nuclear antigen (PCNA), and Ki-67 were compared by double immunostaining and contrasted with single staining of PCNA and Ki-67 indices in the corresponding histological sections from 12 human pituitary adenomas. In vitro, the BUdR labeling index was positive in six of 12 tumors (range < 0.1–5.1%), 10 of 12 tumors were PCNA-positive (range < 0.1–100%), and Ki-67 was positive in 10 of 12 adenomas (range < 0.1–8%). In vitro, BUdR and Ki-67 gave similar proliferative indices for 10 of 12 adenomas. In vivo, the PCNA labeling index was positive in 12 of 12 adenomas (range 0.9–95%) and Ki-67 was positive in 11 of 12 adenomas (range < 0.1–2%). Tumors with a labeling index less than 0.1% were considered to be negative for proliferation. High PCNA values were found in vitro and in vivo, whereas Ki-67 labeling indices were similar in vitro and in vivo for nine of 12 adenomas. It is concluded that Ki-67 proliferative indices in vivo reflect those found in vitro, at least after 4 days in dispersed culture, but that PCNA overestimates pituitary adenoma proliferation in histological sections as well as in dispersed culture.

Human anterior pituitary adenomas proliferate and express the p53 tumour suppressor gene protein, but it is not known if apoptosis (programmed cell death) occurs. Therefore, the detection of apoptosis was undertaken in tumorous human anterior pituitary tissue and compared with p53 protein expression, tumour type and tumour size. Apoptosis (detected by the in situ end labelling technique) and p53 suppressor gene protein (detected by DO. 1-antibody immunocytochemistry) were determined in formalin-fixed and paraffin-embedded tissue from 37 human pituitary adenomas (2 macroprolactinomas, 9 somatotrophinomas and 26 non-functioning adenomas). Two normal anterior pituitaries were also included in this study. Pre-operative tumour size was scored 1 to 4 from magnetic resonance imaging radiology. Apoptosis was found in 7 of 29 tumours (24%), 11% of somatotrophinomas and 33% of non-functioning adenomas, although this difference was not significant. The p53 tumour suppressor protein was found in 7 of 31 tumours (23%), 33% of somatotrophinomas and 19% of nonfunctioning adenomas. Apoptosis and p53 protein expression were not found in normal anterior pituitary. In conclusion, apoptosis occurs in human anterior pituitary adenomas, but no significant association was found between apoptosis and p53 protein expression, tumour type or tumour size. European Journal of Endocrinology 136 382–387

A 14-year-old girl presented with short stature and progressive virilization. She had not undergone the menarche. On investigation, she had elevated testosterone, androstenedione, dihydroepiandrosterone sulphate and 17 alpha-hydroxyprogesterone levels, which were all suppressed by overnight dexamethasone to within their normal ranges. An initial diagnosis of 21-hydroxylase deficiency was revised to 11 beta-hydroxylase deficiency after a tetracosactrin stimulation test, which showed only a modest rise in 17 alpha-hydroxyprogesterone level (from 92 nmol/l at baseline to 133 nmol/l at 60 minutes) and measurement of the basal 11-deoxycortisol, which was grossly elevated. Treatment with dexamethasone 0.5 mg nocte resulted in suppression of androgens in the daytime, but not in the evening, particularly androstenedione. Treatment with hydrocortisone 10 mg b.d. failed to suppress testosterone or androstenedione over a 24-hour period. Addition of cyproterone and oestrogen supplements had no effect and significant virilization persisted. Laparoscopic bilateral adrenalectomy was therefore performed as definitive treatment and resulted in remarkable clinical and biochemical improvement. This case illustrates difficulties in correct diagnosis, choice of appropriate steroid regimen and monitoring efficacy of treatment in congenital adrenal hyperplasia due to 11 beta-hydroxylase deficiency. More aggressive management with earlier bilateral adrenalectomy may be appropriate in selected cases.

Using a differential centrifugation method followed by culture in selective medium, we have successfully isolated and maintained individual epithelial and stromal cells from normal (n = 10) and malignant (n = 6) human breast tissue and characterised their phenotype by immunocytochemistry. Further, we have studied expression of the cytokine genes IL-1β, IL-6, IL-8 and TNF-β in each cell fraction by RT-PCR and have compared these results with cytokine gene expression in tissue extracts from which primary cultures were derived. In breast tumours, there was near complete absence of IL-1β in both whole tissue and cell fractions, and in normals it was present in only 3/10 tissue preparations, with increased expression in stromal (6/10) and epithelial (5/10) cell samples. IL-6 was constitutively expressed in all tumour-derived breast tissue samples but down-regulated in tumour cell cultures, with the opposite result in normal breast. Near identical levels of IL-8 expression were found throughout each preparation, irrespective of tissue origin. TNF-β was expressed in all normal tissue samples, in 9/10 epithelial preparations but in only 6/10 stromal preparations. In tumours, TNF-β was associated predominantly with whole tissue or stromal samples, with reduced expression in epithelial preparations. Our data confirm that primary cultures of normal and malignant human breast tissue can be successfully separated into epithelial and mesenchymal cell populations and their phenotype can be maintained in culture for up to 30 days. However, this cellular separation does alter the cytokine profiles; therefore, experimental findings with isolated cells should be treated with a caveat. © 1996 Wiley-Liss, Inc.

OBJECTIVE The human pituitary gland is affected selectively by conditions associated with iron deposition, but the mechanisms for this are unknown. In this study we have determined whether the transferrin receptor, which mediates iron uptake by cells, could be detected immunocytochemically in human pituitary adenomas in vitro. PATIENTS Data were derived from 35 patients undergoing transsphenoidal surgery and included 13 with clinically non-functioning adenomas, 15 with acromegaly, 4 prolactinomas, 2 patients with Cushing’s disease and one patient with Nelson’s syndrome. MEASUREMENTS Transferrin receptor immunopositivity was determined for each adenoma in dispersed cell culture using a specific monoclonal antibody. RESULTS Eight of 13 clinically functionless adenomas showed immunopositive transferrin receptor expression, whilst adenomas from 15 patients with acromegaly, 4 prolactinomas, 2 Cushing’s syndrome and one patient with Nelson’s syndrome were negative. The eight transferrin receptor positive tumours were gonadotrophinomas and accounted for eight of the nine tumours which secreted and immunostained for FSH; all eight also secreted and immunostained for LH. CONCLUSIONS These findings may reflect a special requirement for iron by gonadotrophin secreting cells in comparison to other pituitary cell types and this could underlie the reasons why in the normal pituitary these cells are especially susceptible to malfunction in iron overload syndromes such as genetic haemochromatosis and β-thalassaemia.

The expression of cytokine transcripts has been investigated in a series of cultured human meningiomas using reverse transcriptase linked polymerase chain reaction (RT-PCR), which allowed simultaneous analysis of a range of cytokines. The main histological subgroups of meningioma were investigated; these included transitional, fibroblastic, and syncytial as well as atypical meningiomas. Meningiomas from each of the different histological subgroups were subjected to a standard tissue culture regime. Total RNA was extracted from representative cultures and reverse-transcribed to yield cDNA. PCR was performed using oligonucleotide primers designed to detect interleukin (IL)-1 alpha/beta to IL-8, transforming growth factor (TGF)beta 1-3, tumour necrosis factor (TNF)alpha/beta, and interferon (IFN)gamma. Transcripts for IL-3, IL-6, IL-8, and TGF beta 3 were detected in all cultures. Transcripts for the three isomers of TGF beta were expressed in the transitional and fibroblastic meningioma cells. TGF beta 2 and TGF beta 3 transcripts were expressed in the syncytial and TGF beta 1 and TGF beta 3 in the atypical meningioma cells. IL-1 beta transcripts were expressed in fibroblastic and atypical cultures and TNF beta transcripts were expressed in syncytial and transitional cultures only. Transcripts for IL-1 alpha, IL-2, IL-4, IL-5, IL-7, TNF alpha, or IFN gamma were not detected in any of the meningioma cultures. This investigation using cells cultured from a small number of tumours from each of the classic histological subtypes suggests that there is a distinct pattern of cytokine mRNA expression linked with histological classification.

The Airtraq(™) optical laryngoscope became available in paediatric sizes in the UK in May 2008. We conducted a randomised, controlled trial comparing the Airtraq with conventional laryngoscopy during routine anaesthesia in children. We hypothesised that the Airtraq laryngoscope would perform as well as conventional laryngoscopy. Sixty patients (20 infants and 40 children) were recruited. The mean (SD) intubation time using the Airtraq was longer than conventional laryngoscopy overall (47.3 (32.6) vs 26.3 (11.5) s; p=0.002), though the difference was only significant for children (p=0.003) and not for infants (p=0.29). The Airtraq provided a better view of the larynx compared with conventional laryngoscopy (in infants (percentage of glottic opening scores 100 (95-100 [90-100]) vs 77 (50-90 [40-100]), respectively; p=0.001; visual analogue scores for field of view 9.2 (9.2-9.5 [8.2-10.0]) vs 6.8 (5.1-8.0 [4.7-10.0]), respectively; p=0.001). In children, the Airtraq provided a similar view of the larynx (percentage of glottic opening scores 100 (100-100 [40-100]) vs 100 (90-100 [50-100]), respectively; visual analogue scores for field of view 9.2 (8.6-10.0 [7.0-10.0]) vs 9.2 (8.6-10.0 [5.6-10.0]), respectively; both p0.05), compared with conventional laryngoscopy.

We examined the production of interleukin (IL)-6 by human meningioma cells in vitro, and the effects of IL-1 beta and IL-4 on IL-6 production and meningioma cell growth. The histological classification of the tumours studied included transitional, syncytial, fibroblastic and atypical. All 10 meningiomas studied produced IL-6 (range 0.22-7.6 ng/ml/10(6) cells/24 h). Separate addition of IL-1 beta or IL-4 to cultures increased IL-6 production up to ten fold, and two to three fold, respectively. Growth studies with IL-6 indicated that this cytokine significantly increased terminal cell density at a concentration greater than 1 ng/ml in 60% of the meningioma cultures studied. IL-1 beta caused a significant decrease in the terminal cell density in 25% of the meningioma cultures studied whereas IL-4 had a tendency to significantly inhibit growth in 16.6% of the cultures. These data suggest that IL-6 production by meningiomas can be modified by other cytokines and secondly, that IL-6, IL-1 beta and IL-4 can modify growth in vitro and may act as autocrine factors in vivo. By further determining the cytokine profiles within meningiomas and their effects, a better understanding of meningioma growth characteristics may be obtained.

Summary A 51-year-old man had been treated for hyperthyroidism with antithyroid drugs for 8 years. He was then found to have a large pituitary adenoma with biochemical evidence of overproduction of TSH, FSH and alpha-subunit. Subsequent immunocytochemical and tissue culture studies confirmed secretion of these hormones. In addition, the tumour stained for GH and was capable of GH production in vitro. This combination of hormones produced by a pituitary adenoma has not been previously reported.

The effects of human recombinant insulin-like growth factor 1 (IGF-1) on the secretion, viability, and proliferation of dispersed human anterior pituitary adenomas secreting FSH, LH, and alpha-subunit (alpha-su) were examined in vitro over 4 h and 4 days. The acute effect of IGF-1 on secretion over 4 h was examined in four tumors secreting FSH, LH, and alpha-su. IGF-1 (100 nmol/L) reduced LH compared to control (100%) in one tumor (61%, P < 0.01), and three tumors remained unaffected. FSH and alpha-su secretion were insufficient to measure over 4 h. Nine tumors were studied over 4 days; relative to control, IGF-1 (100 nmol/L) increased FSH secretion in all seven tumors secreting FSH (28-266%, P < 0.05) and increased alpha-su secretion in all four tumors studied (36%, 63%, 91%, and 121%, P < 0.05). IGF-1 reduced LH secretion in four/nine tumors (13%, 23%, 32%, and 50%, P < 0.05). Dose response curves (1-100 nmol/L IGF-1) were performed on three tumors cosecreting FSH and LH. Stimulation of FSH was achieved with either 1 or 10 nmol/L IGF-1, a single tumor in which alpha-su was measured showed maximal stimulation at 10 nmol/L IGF-1, and one of three tumors showed LH inhibition with 100 nmol/L IGF-1. In situ viability of attached cells was assessed with fluorescein and propidium iodide in seven tumors. After 4 days' exposure to 100 nmol/L IGF-1, in situ viability was increased in five tumors (range 12-19%, 15 +/- 1.3% SEM, P < 0.05). The effects of IGF-1 on the adenoma cell proliferative S-phase fraction was determined in six tumors after 4 days of treatment using double immunostaining with bromodeoxyuridine incorporation for 1 h. In four/six adenomas that stained positive for bromodeoxyuridine in the controls (1-5.6%), the S-phase fraction was increased by 100 nmol/L IGF-1 [(range 2.1-10.6%, increase 90-220%) (P < 0.05)]. These results show that IGF-1 has differential effects on gonadotropins from human pituitary adenomas, stimulating intact FSH and alpha-su, inhibiting or being without effect on intact LH in vitro, and increasing both viability and number of tumorous glycoprotein-secreting cells entering into the S-phase of proliferation.

A new paediatric Glidescope (Cobalt GVL Stat) has recently become available. This varies in design from the Karl Storz DCI videolaryngoscope, as it possesses a short curved disposable blade compared with the narrower straighter blade of the Storz. We compared the time taken for tracheal intubation under normal and difficult intubation conditions in a paediatric manikin. A total of 32 anaesthetists completed four intubations in a random order, with each participant blinded to the airway condition. We hypothesised there would be no difference between the devices. The results showed no difference in tracheal intubation time between the Glidescope and the Storz videolaryngoscope. The mean (SD) times under normal conditions were 18.8 (5.2) s vs 19.9 (6.1) s, (p = 0.16), respectively. Under difficult conditions the times were 22.6 (10.5) vs 27.0 (14.2) s, (p = 0.13), respectively. There were no differences in the visual analogue scores for field of view, ease of use, willingness to use in an emergency, and overall satisfaction.

In this paper we consider the problem of characterizing the variation of the spectrum of a holomorphic family of compact operators ƒ:G → KB(X), where G is an open subset of ℂ and X is a Banach space. The natural conjecture, which the author first heard in a lecture by Professor B. Aupetit, is that these spectra are characterized as those analytic multivalued functions which have as values null sequences. This is obviously a necessary condition, and we prove that this is also sufficient. It will be convenient to use the notation K(ℂ) for the set of compact non-empty subsets of the plane and K0(ℂ) for the subset of K(ℂ) consisting of null sequences.

Serum concentrations of LH and FSH and their response to the separate administration of GnRH (100 micrograms i.v.) and TRH (200 micrograms i.v.) have been studied preoperatively in 12 patients with a clinically functionless pituitary adenoma, of whom nine (3F: 6M) were found to secrete gonadotrophins in vitro. In three patients with a gonadotrophin-secreting adenoma (GSA) the pulsatile release of LH and FSH was also assessed preoperatively. An elevated serum FSH was recorded in six of nine patients with a GSA, and was subnormal in one, whilst an elevated LH was recorded in only two and was subnormal in six. A doubling of LH occurred in only four of the nine patients after GnRH and in three of six after TRH. None of the three patients with a non-GSA was shown to have an aberrant response to GnRH or TRH. In patients with a GSA, pulsatile release of LH and FSH was usually asynchronous and neither hormone demonstrated any regular harmonic pattern. These data show that in patients with a GSA the serum FSH level is usually elevated but this is not invariable, and the LH may well be low. Pathological responses of LH are frequently found following the administration of either GnRH or TRH and these stimulation tests should be performed separately in patients presenting with a clinically 'non-functioning' pituitary tumour to assist in the preoperative diagnosis. The absence of normal LH and FSH pulsing also appears to be a feature of GS adenomas, and suggests that tumorous gonadotrophin secretion is not under physiological control by hypothalamic GnRH.

Spinocerebellar ataxia type 10 (SCA10) is an autosomal dominant disease caused by an expansion of an intronic ATTCT repeat in the ATXN10 gene on 22q13.3.1 Pathogenic alleles have 800 to 4,500 ATTCTs (normal alleles 10 to 29).1,2 Whereas Mexican SCA10 families showed progressive ataxia associated with seizures, polyneuropathy, pyramidal signs, and cognitive and neuropsychiatric impairment,3,4 Brazilian SCA10 families uniformly showed pure cerebellar ataxia.5 We describe clinical and molecular findings in two affected members from an Argentinean family with SCA10. ### Methods. With informed consent, we evaluated three siblings and their mother from an Argentine family with Spanish and Amerindian ancestries (figure, A). The pedigree was extended to deceased members based on the information obtained from the relatives. The analysis of the ATTCT repeated region in the SCA10 gene was performed by PCR, repeat-primed PCR, and Southern blot analyses as described elsewhere.1 Figure The Argentinean family with spinocerebellar ataxia type 10 (SCA10) (A) Pedigree of the family. A number in brackets is the age at onset. Two numbers separated by a diagonal line in parentheses are ATTCT repeat numbers. An arrow …

Twenty-seven prisons in Thailand.To measure the prevalence of smear-positive tuberculosis (TB) and to compare its detection using the World Health Organization (WHO) guidelines for prisons or the International Standards for Tuberculosis Care (ISTC) criteria.Prisoners who were not on TB medications and who had not been treated for TB in the previous 6 months were enrolled in the survey. Prison nurses conducted a one-time mass screening using the WHO questionnaire, with follow-up sputum collection in TB suspects.Of 71594 prisoners, 22132 (30.9%) were identified as TB suspects, and 254 were confirmed by sputum smear, for a prevalence of 354.8/100000. Males were most likely to be diagnosed with TB disease (OR 2.6), as were those transferring in from another prison versus new prisoners (OR 3.0). The positive predictive value of the WHO criteria was 1.2%, and that of the ISTC standard was 5.9%.TB prevalence in this survey remained high. WHO guidelines, recognised as a standard, require health workers to question prisoners and measure height and weight. The ISTC standard, a single question about coughor=2 weeks, is simple, may be asked by non-health personnel and may be a better alternative.

An examination of the reproducibility of three tests for identifying patients at risk of diabetic foot complications is reported. One thousand and one patients attending a general diabetic outpatient clinic at the Royal Liverpool University Hospital underwent screening using the Semmes-Weinstein monofilaments, the biothesiometer, and palpation of pedal pulses. Two hundred and fifty-nine patients who were found to have a deficit on at least one of these tests were given a second appointment where the tests were repeated. Two hundred and twenty-nine patients attended for their second appointment. The expected range of variation in readings for the biothesiometer was 5 log hertz, sufficient variation to make the biothesiometer unacceptable as a screening instrument. The range of reproducibility for the four pedal pulses was from 68% to 81%, also an unacceptable level of variation. Over 85% of the results for the Semmes-Weinstein monofilaments were the same on the first and second measurement. Of the three instruments examined in this study, only the Semmes-Weinstein monofilaments have sufficient reproducibility in the clinical setting for them to be recommended for general use as a screening test for risk of diabetic foot complications.

San Francisco TB Clinic instituted a directly observed preventive therapy (DOPT) program for treatment of persons with latent TB infection in 1996.To examine therapy completion for latent tuberculosis infection before and after implementation of the DOPT program.Medical records were analyzed for patients at the San Francisco TB Clinic referred from high-risk sites for the periods 1993-1994 (n = 619) and 1997-June 1998 (n = 460). Treatment completion and time of therapy were analyzed comparing DOPT to self-administered therapy (SAT).More DOPT patients completed treatment (70.3%) than SAT patients (47.9%) (P0.001). Controlling for sex, age, race/ethnic group and cohort, patients on DOPT were nearly twice as likely to complete therapy (OR 1.93, 95% CI 1.25-3.00).DOPT is a successful strategy. Combined with targeted testing, DOPT can have an important impact in areas with traditionally low rates of treatment adherence.

Minimizing loss to follow-up in longitudinal studies is critical. The purpose of this study was to examine the ability to locate subjects recently released from jail, identify predictors of being able to find a subject, and describe effective search strategies for this unique population. The sample for this cohort study included study subjects who were sought for interview after release from jail. Inmates in the San Francisco City and County Jail were enrolled in a randomized trial of incentives to improve follow-up for tuberculosis therapy after release from jail. Sociodemographic, health-related, and extensive locating information was collected during baseline interviews in jail. The main outcome was successful location of the subject. Study personnel recorded data on the number and nature of attempts made to find subjects in order to describe successful search strategies. Of 254 persons sought for the postrelease interview, 188 (74.0%) were found. Primary English speakers were more likely than Spanish speakers to be found (relative risk: 3.2, 95% confidence interval: 1.5-6.7, p = 0.002). Nearly one quarter of subjects (24%) were found back in jail, and the remainder were found in the community. Phone calls and letters to the subjects, and personal contacts to family and friends were successful strategies for 53% of the subjects. Seeking persons in programs, such as shelters and drug and alcohol programs, was successful in finding 18% of English-speaking subjects. Outreach efforts in sections of the city where Latinos spent time, including popular restaurants and community gathering places, were successful in finding 13% of Spanish-speaking subjects. We conclude that study subjects released from jails can be successfully located using well-defined search protocols tailored to the ethnicity of the sample and including a variety of strategies. Employment of bilingual personnel is important when a large proportion of subjects is monolingual and non-English speaking.

Despite a continuing decline in tuberculosis (TB) in the US, jails remain a high-risk setting for the identification of active and latent TB infection (LTBI).The purpose of this study was to document the change in TB prevalence in the San Francisco City and County Jail.Two period prevalence analyses were done, for 1994 and 1998. The sample included all persons booked into jail during the two years. The rates of inmates screened and the prevalence of active TB and LTBI by sex and ethnicity were compared using computerized records.Prevalence of active TB was 72.1 per 100000 jail population for 1998, and did not change significantly from 1994. In 1998 one third of active TB cases were found through jail screening. Latinos represented respectively 20.1% and 17.7% of those booked in 1994 and 1998, but 43.0% and 41.7% of inmates with LTBI. In 1998, being Latino (odds ratio 2.9) and male (odds ratio 1.6) were most strongly associated with LTBI.Screening for TB among jail inmates is an increasingly valuable clinical and epidemiological tool for case-finding and for identifying persons who would benefit from preventive therapy.

The structural specificity of vitamin D derivatization by PTAD (4-phenyl-1,2,4-triazoline-3,5-dione) was probed using synthetic analogues and ion trap mass spectrometry. EB 1089, a vitamin D(3) analogue which contains a second site for Diels--Alder cycloaddition on its side-chain, allowed the examination of derivatization modes and comparisons of ion fragment structures. The origins of a PTAD-vitamin D(3) ion fragment, commonly used in metabolite characterization and quantitation of vitamin D(3) analogues (m/z 314), were established; ion trap mass spectrometry revealed that the PTAD comprises a portion of this diagnostic fragment, and is not lost by a retro-Diels--Alder step. Furthermore, the unique structure of the EB 1089 side-chain also permits facile determination of its side-chain metabolism. Use of PTAD derivatization and detection of metabolite-specific ion fragments identify hydroxylation at the end of the EB 1089 sidechain. It is believed that the results from these studies provide a clearer understanding of the mass spectrometry of triazolinedione derivatives, not only in the specific case of EB 1089, but also in their application to other vitamin D compounds.

The laboratory investigation of catecholaminesecreting tumours has usually involved measurement of the renal excretion of the catecholamine metabolites, 4-hydroxy-3-methoxymandelic acid (HMMA)1 and/or total metanephrines.i Urinary HMMA measurement, however, has been suggested as unreliable with several reports of cases of excretions within reference limits.v' The estimation of urinary total metanephrines, which were elevated in most of the examples cited, was therefore regarded as the most reliable laboratory index. We now present a case of an adrenalinesecreting tumour where the urinary HMMA excretion was markedly raised compared with the metanephrine excretion. If the latter investigation had been the only one available the laboratory might well have missed the diagnosis.

1alpha,25-dihydroxyvitamin D(3) and two synthetic analogs, 1alpha, 25-dihydroxy-16-ene-23-yne-vitamin D(3) (Ro 23-7553) and 1alpha, 25-dihydroxy-16-ene-24-oxo-vitamin D(3) (JK-1624-3), were tested for their ability to specifically inhibit growth and promote differentiation of human colon cancer cells in comparison with a series of 1beta-(hydroxymethyl) congeners of the natural hormone, such as 1beta-(hydroxymethyl)-3alpha,25(OH)(2)-16-ene,24-oxo-vitamin D(3) (JK-1624-2), 1beta-(hydroxymethyl)-3alpha, 25-dihydroxy-16-ene-26,27-dihomo vitamin D(3) (JK-1626-2), and 1beta-(hydroxymethyl)-3alpha,25-dihydroxy-22,24-diene-26,27- dihomo vitamin D(3) (MCW-EE). Western blot analysis revealed that reduction of cyclin D1 levels is a key mechanism by which the vitamin D compounds under investigation inhibit Caco-2 tumor cell growth. Both the 1alpha-hydroxy- as well as the 1beta-hydroxymethyl-type vitamin D compounds, which exhibit only low affinity for the vitamin D receptor, significantly reduced [(3)H]thymidine DNA labeling in confluent Caco-2 cell cultures. This suggests that high-affinity binding to the vitamin D receptor is not an absolute prerequisite for genomic action on tumor cell growth. Hybrid analogs JK-1624-2 and MCW-EE, although antimitotically active, were rather ineffective in promoting phenotypic differentiation of human colon cancer cells. However, because both compounds also do not promote osteoclast differentiation from hematopoetic bone marrow cells, they still could be used as antimitotic agents in cancer therapy, even at dose levels that, with other analogs, could cause hypercalcemia.

The presence of mRNA transcripts for cytokines in normal and neoplastic human breast tissue has been investigated. Using reverse transcriptase-linked polymerase chain reaction (RT-PCR), we have specifically screened for the following cytokines: interleukin (IL)-1alpha, IL-1beta, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, tumour necrosis factor (TNF)-alpha, TNF-beta and interferon (IFN)-gamma. No significant differences in expression of IL-1alpha, IL-1beta, IL-4, IL-6, TNF-alpha or TNF-beta were observed between the 2 groups of tissues. However, there was a significant difference in expression of IL-8 transcripts (p = 0.0017) which was higher in the neoplastic population. Transcripts for IL-2, IL-3, IL-5, IL-7 and IFN-gamma were not detected in either group. There was no evidence of associations between cytokine expression and tumour histological grade, patient age or lymph node metastases. Correlating tumour types with specific cytokine transcripts revealed high expression of IL-8, and to a lesser extent, IL-8 and TNF-beta irrespective of tumour origin. Analysis of primary epithelial and stromal cultures derived from both types of tissue showed that increased levels of IL-8, but not IL-6, were secreted by cells obtained from tumours. Thus, breast tissue of both normal and neoplastic origin expresses a wide range of cytokines. Increased or aberrant expression of cytokines, in particular IL-8, may be involved in the development/progression of breast cancer.

1,25-(OH)2D3 and 24,25-(OH)2D3 mediate their effects on chondrocytes through the classic vitamin D receptor (VDR) as well as through rapid membrane-mediated mechanisms which result in both nongenomic and genomic effects. In intact cells, it is difficult to distinguish between genomic responses via the VDR and genomic and nongenomic responses via membrane-mediated pathways. In this study, we used two hybrid analogues of 1,25-(OH)2D3 which have been modified on the A-ring and C,D-ring side chain (1 alpha-(hydroxymethyl)-3 beta-hydroxy-20-epi-22-oxa-26,27-dihomo vitamin D3 (analogue MCW-YA = 3a) and 1 beta-(hydroxymethyl)-3 alpha-hydroxy-20-epi-22-oxa-26,27-dihomo vitamin D3 (analogue MCW-YB = 3b) to examine the role of the VDR in response of rat costochondral resting zone (RC) and growth zone (GC) chondrocytes to 1,25-(OH)2D3 and 24,25-(OH)2D3. These hybrid analogues are only 0.1% as effective in binding to the VDR from calf thymus as 1,25-(OH)2D3. Chondrocyte proliferation ([3H]-thymidine incorporation), proteoglycan production ([35S]-sulfate incorporation), and activity of protein kinase C (PKC) were measured after treatment with 1,25-(OH)2D3, 24,25-(OH)2D3, or the analogues. Both analogues inhibited proliferation of both cell types, as did 1,25-(OH)2D3 and 24,25-(OH)2D3. Analogue 3a had no effect on proteoglycan production by GCs but increased that by RCs. Analogue 3b increased proteoglycan production in both GC and RC cultures. Both analogues stimulated PKC in GC cells; however, neither 3a nor 3b had an effect on PKC activity in RC cells. 1,25-(OH)2D3 and 3a decreased PKC in matrix vesicles from GC cultures, whereas plasma membrane PKC activity was increased, with 1,25-(OH)2D3 having a greater effect. 24,25-(OH)2D3 caused a significant decrease in PKC activity in matrix vesicles from RC cultures; 24,25-(OH)2D3, 3a, and 3b increased PKC activity in the plasma membrane fraction, however. Thus, with little or no binding to calf thymus VDR, 3a and 3b can affect cell proliferation, proteoglycan production, and PKC activity. The direct membrane effect is analogue-specific and cell maturation-dependent. By studying analogues with greatly reduced affinity for the VDR, we have provided further evidence for the existence of a membrane receptor(s) involved in mediating nongenomic effects of vitamin D metabolites.

The purpose of this study was to describe the perceived health of the homeless, and to measure the effect of time homeless on perceived health status, after controlling for sociodemographic characteristics and health conditions. The design was cross-sectional; the population was a representative sample of homeless in San Francisco, interviewed on health issues. Analysis of predictors of poor or fair health status was by logistic regression. In this sample of 2780 persons, 37.4% reported that their health status was poor or fair as compared to good or excellent. Reporting poor or fair health status was significantly associated with time homeless, after controlling for sociodemographic variables and health problems including results from screening for HIV and TB (OR = 1.49, 95% CI 1.24-1.79). Comparisons with data from the National Health Interview Survey (NHIS) showed poorer health status among the homeless persons in this study. Standardized morbidity ratios were highest for asthma; there was twice the number of homeless persons reporting asthma, in younger as well as older adults, as would be expected using NHIS rates. There was also an excess of arthritis, high blood pressure and diabetes in those age 18-44 as compared to adults in the Health Interview Survey. The time spent homeless remains associated with self-reported health status, after known contributors to poor health are controlled. Persons who have been homeless for longer periods of time may be the persons to whom health care interventions should be aimed.

An autolysis-resistant mutant of the HIV-I protease was employed for removal of metabolically stabilized and highly bioactive analogues of bovine growth-hormone-releasing factor (bGRF) from their larger either synthetic or recombinant precursors. The N-terminal four amino acids in two selected model GRF analogues, Y1IDAIFTSSYRKVLAQLSARKLLQDILSRQVF32-OH (I; GRF32) and Y1IDAIFTSSYRKVLAQLSARKLLQDILSRQ30-OH (IA; GRF30), conform well to the specificity of the HIV-I protease for residues in the P1' to P4' positions of its peptide substrates. A variety of amino acids were tried in the N-terminal extension (positions P4-P1) to fit the protease substrate specificity for the 8 amino acids in positions P4-P4'. A synthetic precursor of I, extended N-terminally with RQVF-, a sequence representing the four C-terminal residues in I, was effectively cleaved by the protease at the Phe-1-Tyr1 bond (... RQVF-decreases-YIDA ...) to release GRF32. However, when several soluble fusion proteins linked to GRF32 by the RQVF sequence were expressed in Escherichia coli, attempts to cleave out the core GRF32 met with variable, and only limited, success. By random mutagenesis in a propeptide segment, [MGQSVAQVF]-decreases-GRF30, (II) was identified as a construct that showed reasonably high-level expression in E. coli and was effectively processed by the HIV-I protease. A yield of 5 mg of pure GRF30 was obtained/litre of culture medium after a single HPLC purification step.

The combination of 10-12 kbar pressure plus Lewis acidic zinc dichloride promotes highly regioselective and stereoselective Diels-Alder cycloaddition between 3-bromo-2-pyrone, prepared in a new way, and unactivated terminal alkene 4-(tert-butyldimethylsiloxy)-1-butene. The conjugate base of A-ring allylic phosphine oxide 20 adds chemospecifically to the C-8 keto group of some C-8,C-24-diketones to form directly metabolically resistant 24-oxo analogs of 1alpha,25-dihydroxyvitamin D(3) (calcitriol). Several of these new hybrid analogs are as efficacious in vitro as calcitriol at inhibiting growth of murine keratinocytes even at physiologically relevant 10-100 nanomolar concentrations.

1 alpha-hydroxymethyl-25-hydroxyvitamin D3 and 1 beta-hydroxymethyl -3 alpha,25-hydroxyvitamin D3, two analogs with modifications restricted to the A ring, bind poorly to vitamin D receptor (VDR). The effective doses required for 50% of maximal binding activity (ED50) are 7 x 10(-7) M for the former and 8 x 10(-8) M for the latter, and the ED50 for their growth-inhibitory activities is greater than 10(-6) M. Unexpectedly, a hybrid analog with 20-epi configuration at its side chain and a 1 beta-hydroxymethyl group but not a 1 alpha-hydroxymethyl group inhibits malignant cell growth with an ED50 of 7 x 10(-9) M. To determine if the restored biological activity of the hybrid analog is associated with better binding to VDR, we performed competitive binding assays in vitro with calf thymus VDR and in vivo with recombinant human VDR. We found that the 20 epi side chain reduced the affinity of the 1 beta- and the 1 alpha-hydroxymethyl hybrid analogs for VDR in vitro and in vivo fourfold to tenfold. To determine whether the 1 beta-hydroxymethyl analogs induced a VDR-mediated transcription, we tested the induction of reporter gene expression through the osteocalcin vitamin D response element (VDRE) in ROS 17/2.8 cells and the induction of binding activity of VDR to VDRE in COS-1 cells. We found that the ED50 for transcriptional activity of 1 beta-hydroxymethyl-3 alpha,25-hydroxyvitamin D3 was greater than 10(-6) M, but its 1 alpha diastereomer had barely detectable transcriptional activity. The 20-epi side chain preferentially increased the transcriptional activity of the 1 beta-hydroxymethyl hybrid analog to an ED50 of 10(-8) M, but the 1 alpha-hydroxymethyl hybrid analog remained inactive. To confirm that this transcriptional activity was dependent on the VDR, we repeated the assay in VDR-negative CV-1 cells and compared ligand-dependent expression of the VDRE/growth hormone reporter in the presence of either wild-type or transcriptionally inactive mutant VDR expression vectors. Transcription was induced by the 1 beta-hydroxymethyl compounds only in the presence of wild-type VDR. Thus, we conclude that it is possible, by adding a 20 epi side chain, to restore growth-inhibitory and VDR-mediated transcriptional activities without increasing binding to the VDR of A ring-modified analogs.

Atkin SL, Coady AM. White MC, Mathew B. Hyponatraemia secondary to cerebral salt wasting syndrome following routine pituitary surgery. Eur J Endocrinol 1996;135:245–7. ISSN 0804–4643 A female aged 53 years was found to have a suprasellar lesion, which was shown to be a Rathke's cyst after removal by transsphenoidal surgery. She presented 16 days postoperatively, and following two grand mal seizures was found to be profoundly hyponatraemic (sodium 101 mmol/l). She was initially thought to have the syndrome of inappropriate antidiuretic hormone and was treated accordingly, but central venous pressure measurement revealed the hypovolaemia of cerebral salt wasting syndrome. The patient subsequently developed severe neurological sequelae after the correction of her hyponatraemia, following the development of extrapontine myelinolysis. Cerebral salt wasting syndrome is a rare cause of hyponatraemia following pituitary transsphenoidal surgery, which may mimic the syndrome of inappropriate antidiuretic hormone secretion. This case emphasizes the poor prognosis that may result from the rapid correction of profound hyponatraemia. SL Atkin, Michael White Centre for Diabetes and Endocrinology, Royal Hull Hospitals, 220-236 Anlaby Road, Hull HU3 2RW, UK

Using a differential centrifugation method followed by culture in selective medium, we have successfully isolated and maintained individual epithelial and stromal cells from normal (n = 10) and malignant (n = 6) human breast tissue and characterised their phenotype by immunocytochemistry. Further, we have studied expression of the cytokine genes IL-1beta, IL-6, IL-8 and TNF-beta in each cell fraction by RT-PCR and have compared these results with cytokine gene expression in tissue extracts from which primary cultures were derived. In breast tumours, there was near complete absence of IL-1beta in both whole tissue and cell fractions, and in normals it was present in only 3/10 tissue preparations, with increased expression in stromal (6/10) and epithelial (5/10) cell samples. IL-6 was constitutively expressed in all tumour-derived breast tissue samples but down-regulated in tumour cell cultures, with the opposite result in normal breast. Near identical levels of IL-8 expression were found throughout each preparation, irrespective of tissue origin. TNF-beta was expressed in all normal tissue samples, in 9/10 epithelial preparations but in only 6/10 stromal preparations. In tumours, TNF-beta was associated predominantly with whole tissue or stromal samples, with reduced expression in epithelial preparations. Our data confirm that primary cultures of normal and malignant human breast tissue can be successfully separated into epithelial and mesenchymal cell populations and their phenotype can be maintained in culture for up to 30 days. However, this cellular separation does alter the cytokine profiles; therefore, experimental findings with isolated cells should be treated with a caveat.