Chemosensory behaviour of juvenile crown-of-thorns sea star (Acanthaster sp.), attraction to algal and coral food and avoidance of adult conspecifics.

Intraspecific and habitat-mediated responses to chemical cues play key roles in structuring populations of marine species. We investigated the behaviour of herbivorous-stage juvenile crown-of-thorns sea stars (COTS; Acanthaster sp.) in flow-through choice chambers to determine if chemical cues from their habitat influence movement and their transition to become coral predators. Juveniles at the diet transition stage were exposed to cues from their nursery habitat (coral rubble-crustose coralline algae (CCA)), live coral and adult COTS to determine if waterborne cues influence movement. In response to CCA and coral as sole cues, juveniles moved towards the cue source and when these cues were presented in combination, they exhibited a preference for coral. Juveniles moved away from adult COTS cues. Exposure to food cues (coral, CCA) in the presence of adult cues resulted in variable responses. Our results suggest a feedback mechanism whereby juvenile behaviour is mediated by adult chemical cues. Cues from the adult population may deter juveniles from the switch to corallivory. As outbreaks wane, juveniles released from competition may serve as a proximate source of outbreaks, supporting the juveniles-in-waiting hypothesis. The accumulation of juveniles within the reef infrastructure is an underappreciated potential source of COTS outbreaks that devastate coral reefs.

Intrathecal non-viral interleukin-10 gene therapy ameliorates neuropathic pain as measured by both classical static allodynia and a novel supra-spinally mediated pain assay, the Two-Arm Rodent Somatosensory (TARS) task.

Intrathecal delivery of interleukin-10 (IL-10) gene therapy has been reported to be effective in suppressing pain enhancement in a variety of rodent models. However, all publications that have tested this treatment have relied upon measures of static allodynia (von Frey test) and thermal hyperalgesia (Hargreaves test). As this plasmid DNA IL-10 (pDNA-IL10) therapeutic approach is now in human clinical trials for multiple pain indications, including intrathecal delivery for human neuropathic pain, it is important to consider the recent concerns raised in the pain field that such tests reflect spinal rather than supraspinal processing of, and responsivity to, noxious stimuli. Consequently, this raises the question of whether intrathecal pDNA-IL10 can reverse established neuropathic pain when assessed by a test requiring supraspinal, rather than solely spinal, mediation of the behavioral response. The present study utilizes the rat sciatic chronic constriction injury (CCI) model of neuropathic pain to compare the expression of static allodynia with that of cognitively controlled choice behavior in a two-arm maze, adapted from Hayashida et al. (2019). This modification, termed the Two-Arm Rodent Somatosensory (TARS) task, provides rats free choice to reach a desired goal box via a short "arm" of the maze with tactile probes as flooring versus a longer "arm" of the maze with a smooth surface. Here we demonstrate that static allodynia and avoidance of the nociceptive flooring in TARS develop in parallel over time, and that both behaviors also resolve in parallel following intrathecal pDNA-IL10 gene therapy. Details for the construction and use of this new maze design are also provided. Together, this study documents both: (a) the important finding that intrathecal IL-10 gene therapy does indeed resolve neuropathic pain as measured by a supraspinally-mediated behavioral task, and (b) a new, supraspinally-mediated task that allows behavioral assessments across weeks and allows the analysis of both development and resolution of neuropathic pain by therapeutic interventions. As such, the TARS operant behavior task is an improvement over other approaches such as the mechanical conflict-avoidance system which have difficulties demonstrating development and reversal of pain behavior in a within-subject design.

The Transatlantic HbA1c gap: differences in glycaemic control across the lifespan between people included in the US T1D Exchange Registry and those included in the German/Austrian DPV registry.

AIM: To compare HbA1c levels across the lifespan in people with type 1 diabetes in the USA with those in Germany/Austria, and to examine potential differences in HbA1c levels between sexes, insulin delivery methods and minority status. METHODS: Data were extracted from the US T1D Exchange Registry (n=18 381 participants from 73 sites) and from the German/Austrian Prospective Diabetes Follow-up Registry, the DPV (n=32 643 participants from 362 sites). Mean HbA1c was calculated for each year of age for individuals aged ≤25 years, and at 2-year age intervals for individuals aged >25 years. Curves for mean HbA1c by age were estimated using locally weighted scatterplot smoothing. HbA1c differences between registries, sexes, insulin delivery methods, and minority status were assessed by age group using multiple linear regression. RESULTS: In both registries, mean HbA1c increased by ~11 mmol/mol (1.0%) between the ages of 9 and 18 years, although at quite different absolute levels: from 66 mmol/mol (8.2%) to 77 mmol/mol (9.2%) in the T1D Exchange Registry, and from 56 mmol/mol (7.3%) to 66 mmol/mol (8.2%) in the DPV. Sex differences were observed in the DPV only. In the T1D Exchange Registry, injection users had higher mean HbA1c than pump users across the lifespan, whereas in the DPV higher HbA1c levels in injection users were observed in the age groups 6 to <12 years, 12 to <18 years, and 30 to <50 years (P < 0.001). Minority status was significantly associated with higher HbA1c in most age groups in both registries. CONCLUSIONS: Significant differences in HbA1c were noted between the USA and Germany/Austria, with disparities more pronounced in early childhood through to young adulthood. Further studies should identify causes for these disparities.

Long-term excess risk of stroke in people with Type 2 diabetes in Sweden according to blood pressure level: a population-based case-control study.

AIMS: To estimate the risk of stroke in people with Type 2 diabetes with different blood pressure levels compared with the risk in the general population in Sweden. METHODS: This prospective case-control study included 408 076 people with Type 2 diabetes, aged ≥ 18 years, and free of prior stroke, registered in the Swedish National Diabetes Register 1998-2011. Age- and sex-matched control subjects (n = 1 913 507) without stroke from the general population were included. Stroke diagnoses were retrieved using International Classification of Disease codes from the Swedish patient and death registers. Cox hazard ratios and 95% confidence intervals (CIs) were estimated at six different blood pressure levels. RESULTS: During a median follow-up of 4 years, 19 548 (4.8%) people with Type 2 diabetes and 61 690 (3.2%) without diabetes were diagnosed with stroke, corresponding to an adjusted hazard ratio of 1.43 (95% CI 1.41-1.46) for people with Type 2 diabetes as a group. Compared with people without diabetes, the risk of stroke for people with Type 2 diabetes with different blood pressure levels was significantly higher, starting at blood pressure levels > 130/80 mmHg. Hazard ratios for stroke were 1.20 (95% CI 1.16-1.24), 1.47 (95% CI 1.43-1.50), and 1.97 (95% CI 1.90-2.03) for blood pressure categories of 130-139/80-89 mmHg, 140-159/90-99 mmHg and ≥ 160/≥ 100 mmHg, respectively, after adjustment for age, sex, diabetes duration, being born in Sweden, maximum education level and baseline comorbidities. CONCLUSIONS: People with Type 2 diabetes and blood pressure < 130/80 mmHg had a risk of stroke similar to that of the general population.

RT-Bst: an integrated approach for reverse transcription and enrichment of cDNA from viral RNA.

The synthesis of cDNA from RNA is challenging due to the inefficiency of reverse transcription (RT). In order to address this, an RT-Bst method was developed for sequential RT of RNA and Bst DNA polymerase amplification for enrichment of cDNA in a single-tube reaction. Using genomic RNA from bacteriophage MS2, the yield of cDNA produced by RT alone and RT-Bst were compared by analysis of polymerase chain reaction (PCR)-amplified products. A superior performance was observed when amplifying MS2 cDNA with random primers following RT-Bst compared to RT alone, indicating greater quantities of cDNA were present after RT-Bst. RT-Bst was also compared with RT alone for their relative ability to produce sufficient cDNA to amplify eight target regions spanning the respiratory syncytial virus (RSV) genome. Six out of eight targets were amplified consistently by PCR subsequent to RT-Bst amplification, whereas only three out of eight targets could be amplified after RT alone. The RSV sequences were selectively amplified using RSV-specific primers from a mixed template containing an excess of MS2 RNA without amplifying MS2 sequences. This suggests that RT-Bst can be used to amplify RNA sequences non-specifically using random primers and specifically using sequence-specific primers, and enhances the yield of cDNA when compared to RT alone.

Application of RT-Bst to enhance detection of pathogenic viruses of the respiratory tract.

Inefficiency of RT-PCR can be associated with the suboptimal process of reverse transcription as only 40-80% of RNA is converted to cDNA. We employed a novel method, RT-Bst, to enrich the concentration of cDNA for subsequent multiplex PCR detection of selected RNA viruses. The RT-Bst method amplifies cDNA through reverse transcription of viral RNA using reverse transcriptase and amplification of cDNA using Bst DNA polymerase. Viral RNA was extracted from 25 nasopharyngeal samples for detection of influenza A, B and C; parainfluenza 1-4; human coronaviruses 229E and OC43; respiratory syncytial virus (RSV) and rhinovirus. Both multiplex one-step RT-PCR and RT-Bst PCR were used to compare their performances for detection of virus sequences. These findings were compared with routine laboratory detection. When using RT-Bst PCR, 28% of samples yielded a viral pathogen compared to 20% with RT-PCR and 12% using routine diagnostic tests. RT-Bst PCR was shown to have particular utility in the detection of RSV RNA as this was present in 20% of the samples studied compared to 8% when using RT-PCR. For one patient, RT-Bst PCR was able to detect RSV five days earlier than conventional hospital diagnostic testing. RT-Bst and RT-Bst PCR can be used as alternative approaches to reverse transcription and one-step RT-PCR, respectively, for sequence-independent amplification of RNA virus sequences and a larger scale analysis of this new diagnostic approach is warranted.

Time-dependent risk of developing distant metastasis in breast cancer patients according to treatment, age and tumour characteristics.

BACKGROUND: Metastatic breast cancer is a severe condition without curative treatment. How relative and absolute risk of distant metastasis varies over time since diagnosis, as a function of treatment, age and tumour characteristics, has not been studied in detail. METHODS: A total of 9514 women under the age of 75 when diagnosed with breast cancer in Stockholm and Gotland regions during 1990-2006 were followed up for metastasis (mean follow-up=5.7 years). Time-dependent development of distant metastasis was analysed using flexible parametric survival models and presented as hazard ratio (HR) and cumulative risk. RESULTS: A total of 995 (10.4%) patients developed distant metastasis; the most common sites were skeleton (32.5%) and multiple sites (28.3%). Women younger than 50 years at diagnosis, with lymph node-positive, oestrogen receptor (ER)-negative, >20 mm tumours and treated only locally, had the highest risk of distant metastasis (0-5 years' cumulative risk =0.55; 95% confidence interval (CI): 0.47-0.64). Women older than 50 years at diagnosis, with ER-positive, lymph node-negative and ≤20-mm tumours, had the same and lowest cumulative risk of developing metastasis 0-5 and 5-10 years (cumulative risk=0.03; 95% CI: 0.02-0.04). In the period of 5-10 years after diagnosis, women with ER-positive, lymph node-positive and >20-mm tumours were at highest risk of distant recurrence. Women with ER-negative tumours showed a decline in risk during this period. CONCLUSION: Our data show no support for discontinuation at 5 years of clinical follow-up in breast cancer patients and suggest further investigation on differential clinical follow-up for different subgroups of patients.

Prospective cohort study of body mass index and the risk of hospitalisation: findings from 246361 participants in the 45 and Up Study.

OBJECTIVE: To quantify the risk of hospital admission in relation to fine increments in body mass index (BMI). DESIGN, SETTING, AND PARTICIPANTS: Population-based prospective cohort study of 246,361 individuals aged greater than or equal to 45 years, from New South Wales, Australia, recruited from 2006-2009. Self-reported data on BMI and potential confounding/mediating factors were linked to hospital admission and death data. MAIN OUTCOMES: Cox-models were used to estimate the relative risk (RR) of incident all-cause and diagnosis-specific hospital admission (excluding same day) in relation to BMI. RESULTS: There were 61,583 incident hospitalisations over 479,769 person-years (py) of observation. In men, hospitalisation rates were lowest for BMI 20-<25 kg m(-2) (age-standardised rate: 120/1000 py) and in women for BMI 18.5-<25 kg m(-2) (102/1000 py); above these levels, rates increased steadily with increasing BMI; rates were 203 and 183/1000 py, for men and women with BMI 35-50 kg m(-2), respectively. This pattern was observed regardless of baseline health status, smoking status and physical activity levels. After adjustment, the RRs (95% confidence interval) per 1 kg m(-2) increase in BMI from ≥ 20 kg m(-2) were 1.04(1.03-1.04) for men and 1.04(1.04-1.05) for women aged 45-64; corresponding RRs for ages 65-79 were 1.03(1.02-1.03) and 1.03(1.03-1.04); and for ages ≥ 80 years, 1.01(1.00-1.01) and 1.01(1.01-1.02). Hospitalisation risks were elevated for a large range of diagnoses, including a number of circulatory, digestive, musculoskeletal and respiratory diseases, while being protective for just two-fracture and hernia. CONCLUSIONS: Above normal BMI, the RR of hospitalisation increases with even small increases in BMI, less so in the elderly. Even a small downward shift in BMI, among those who are overweight not just those who are obese, could result in a substantial reduction in the risk of hospitalisation.

Quantitative proteomic analysis of the response of the wood-rot fungus, Schizophyllum commune, to the biocontrol fungus, Trichoderma viride.

AIMS: Investigation of changes in the protein profile of the wood-rot fungus, Schizophyllum commune, when paired against the biocontrol fungus, Trichoderma viride, for 48 h. METHODS AND RESULTS: Variations in protein profile resulting from contact with T. viride were assessed by spot separation using 2 dimensional protein gel electrophoresis followed by MALDI-TOF-TOF MS/MS protein identification. Contact with T. viride elicited a systematic response in S. commune, characterized by marked increases in proteins involved for transcription and translation (61%) and cell wall/hyphal biogenesis and stabilization (17%), whereas metabolism-associated proteins decreased in amounts (64%). Trichoderma viride, however, exhibited typical mycoparasitic behaviour with increases in the amounts of proteins involved in proteolysis and carbohydrate metabolism. CONCLUSIONS: The protein profile of S. commune confronted by T. viride indicates the up-regulation of mechanisms specifically targeted at the mycoparasitic machinery of T. viride, particularly cell wall lysis and antibiosis. SIGNIFICANCE AND IMPACT OF THE STUDY: The proteomic responses observed in S. commune may occur in natural environments, providing an insight to the mechanism involved in conferring resistance to mycoparasitic attack. This study, therefore, warrants further investigation for the targeted design of more robust biocontrol agents.

Comparative clonal analysis of human immunodeficiency virus type 1 (HIV-1)-specific CD4+ and CD8+ cytolytic T lymphocytes isolated from seronegative humans immunized with candidate HIV-1 vaccines.

The lysis of infected host cells by virus-specific cytolytic T lymphocytes (CTL) is an important factor in host resistance to viral infection. An optimal vaccine against human immunodeficiency virus type 1 (HIV-1) would elicit virus-specific CTL as well as neutralizing antibodies. The induction by a vaccine of HIV-1-specific CD8+ CTL in humans has not been previously reported. In this study, CTL responses were evaluated in HIV-1-seronegative human volunteers participating in a phase I acquired immune deficiency syndrome (AIDS) vaccine trial involving a novel vaccine regimen. Volunteers received an initial immunization with a live recombinant vaccinia virus vector carrying the HIV-1 env gene and a subsequent boost with purified env protein. An exceptionally strong env-specific CTL response was detected in one of two vaccine recipients, while modest but significant env-specific CTL activity was present in the second vaccinee. Cloning of the responding CTL gave both CD4+ and CD8+ env-specific CTL clones, permitting a detailed comparison of critical functional properties of these two types of CTL. In particular, the potential antiviral effects of these CTL were evaluated in an in vitro system involving HIV-1 infection of cultures of normal autologous CD4+ lymphoblasts. At extremely low effector-to-target ratios, vaccine-induced CD8+ CTL clones lysed productively infected cells present within these cultures. When tested for lytic activity against target cells expressing the HIV-1 env gene, CD8+ CTL were 3-10-fold more active on a per cell basis than CD4+ CTL. However, when tested against autologous CD4+ lymphoblasts acutely infected with HIV-1, CD4+ clones lysed a much higher fraction of the target cell population than did CD8+ CTL. CD4+ CTL were shown to recognize not only the infected cells within these acutely infected cultures but also noninfected CD4+ T cells that had passively taken up gp120 shed from infected cells and/or free virions. These results were confirmed in studies in which CD4+ lymphoblasts were exposed to recombinant gp120 and used as targets for gp120-specific CD4+ and CD8+ CTL clones. gp120-pulsed, noninfected targets were lysed in an antigen-specific fashion by CD4+ but not CD8+ CTL clones. Taken together, these observations demonstrate that in an in vitro HIV-1 infection, sufficient amounts of gp120 antigen are produced and shed by infected cells to enable uptake by cells that are not yet infected, resulting in the lysis of these noninfected cells by gp120-specific, CD4+ CTL.(ABSTRACT TRUNCATED AT 400 WORDS)

Type and quantity of data needed for an early estimate of transmissibility when an infectious disease emerges.

An early estimate of disease transmissibility is essential for a well-informed public health response to a newly emerged infectious disease. In this study, we ask what type and quantity of data are needed for useful estimation of the initial reproduction number (R). It is possible to estimate R from case incidence data alone when the growing incidence of cases displays a wave pattern, because the pattern provides information about the serial interval (the time elapsed between the onset of symptoms of a case and symptom onset in individuals infected by that case). When the mode of the serial interval distribution is small, 1.5 days or less, there is generally no informative wave pattern in the observed series of daily incidences. The precision of the estimate of R is then improved substantially by having some observations on the serial interval. For an infectious disease with characteristics such as those of influenza, an estimate of R able to inform plans to mitigate transmission is obtained when the cumulative incidence of cases reaches about 300 and about 10 observations on the serial interval are available.

Oral amoxicillin and amoxicillin-clavulanic acid: properties, indications and usage.

BACKGROUND: Amoxicillin has been in use since the 1970s; it is the most widely used penicillin both alone and in combination with the ß-lactamase clavulanic acid. OBJECTIVES: In this narrative review, we re-examine the properties of oral amoxicillin and clavulanic acid and provide guidance on their use, with emphasis on the preferred use of amoxicillin alone. SOURCES: Published medical literature (MEDLINE database via Pubmed). CONTENT: While amoxicillin and clavulanic acid have similar half-lives, clavulanic acid is more protein bound and even less heat stable than amoxicillin, with primarily hepatic metabolism. It is also more strongly associated with gastrointestinal side effects, including Clostridium difficile infection, and, thus, in oral combination formulations, limits the maximum daily dose of amoxicillin that can be given. The first ratio for an amoxicillin-clavulanic acid combination was set at 4:1 due to clavulanic acid's high affinity for ß-lactamases; ratios of 2:1, 7:1, 14:1 and 16:1 are currently available in various regions. Comparative effectiveness data for the different ratios are scarce. Amoxicillin-clavulanic acid is often used as empiric therapy for many of the World Health Organization's Priority Infectious Syndromes in adults and children, leading to extensive consumption, when some of these syndromes could be handled with a delayed antibiotic prescription approach or amoxicillin alone. IMPLICATIONS: Using available epidemiological and pharmacokinetic data, we provide guidance on indications for amoxicillin versus amoxicillin-clavulanic acid and on optimal oral administration, including choice of combination ratio. More data are needed, particularly on heat stability, pharmacodynamic effects and emergence of resistance in 'real-world' clinical settings.

Description of a fertilization-independent obligate apomictic species: Corunastylis apostasioides Fitzg.

The Australian midge orchid Corunastylis apostasioides of the tribe Diurideae has completely eliminated any male contribution in the process of seed formation, which occurs directly from the maternal tissue by a process termed apomixis. Here, we report C. apostasioides to be an obligate apomictic species devoid of any sexuality and compare its development to a close sexual relative C. fimbriata (R. Br.) D.L. Jones & M.A. Clem. Apomictic characteristics in C. apostasioides include production of seed in absence of fertilization, frequently closed flowers, production of immature pollen in non-dehiscent anthers, expansion of ovaries despite the lack of fertilization and the absence of a citronella scent that is found in C. fimbriata produced to attract pollinating vinegar flies (Jones 2006). The nature of apomixis in C. apostasioides was examined by ovule histology and amplified fragment length polymorphism (AFLP) in each case drawing comparison with sexual C. fimbriata. In C. apostasioides the central megaspore mother cell undergoes diplosporic apomixis, while additional embryos are derived from nucellar or integument initials formed by sporophytic apomixis. Typical of apomicts, C. apostasioides is polyploid compared to the sexual C. fimbriata. The divergences of C. apostasioides from sexuality to apomictic development are discussed.

Induction of CD4+ human cytolytic T cells specific for HIV-infected cells by a gp160 subunit vaccine.

Cytolytic T lymphocyte (CTL) responses were evaluated in humans immunized with recombinant human immunodeficiency virus type 1 (HIV) envelope glycoprotein gp160. Some vaccinees had gp160-specific CTLs that were shown by cloning to be CD4+. Although induced by exogenous antigen, most gp160-specific CTL clones also recognized gp160 synthesized endogenously in target cells. These clones lysed autologous CD4+ T lymphoblasts infected with HIV. Of particular interest were certain vaccine-induced clones that lysed HIV-infected cells, recognized gp160 from diverse HIV isolates, and did not participate in "innocent bystander" killing of noninfected CD4+ T cells that had bound gp120.

Improved glycaemic control of thrice-daily biphasic insulin aspart compared with twice-daily biphasic human insulin; a randomized, open-label trial in patients with type 1 or type 2 diabetes.

AIM: This trial evaluated the potential for improving glycaemic control by intensifying a conventional twice-daily therapy with premixed human insulin (HI) to a thrice-daily regimen using premixed formulations of biphasic insulin aspart (BIAsp) in patients with type 1 or type 2 diabetes. METHODS: This was a multicentre, open-label, parallel group trial. After a 4-week run-in period, patients were randomized 1 : 1 to 16 weeks of treatment. A total of 748 patients were screened, 664 were exposed to trial drug and 604 completed the trial. RESULTS: Haemoglobin A(1c), the primary efficacy endpoint, was shown to be significantly lower for the BIAsp treatment group compared with the biphasic HI (BHI) 30 group [estimated mean difference: -0.32, 95% confidence interval (CI) (-0.48; -0.16), p = 0.0001]. The average blood glucose level was significantly lower in the BIAsp group [estimated mean difference: -0.79, 95% CI (-1.17; -0.40), p = 0.0001]. There were few major hypoglycaemic episodes, 11 in the BIAsp group and 7 in the BHI 30 group. Although intensification of insulin therapy with BIAsp three times a day was associated with a higher risk of minor hypoglycaemia (relative risk = 1.58, p = 0.0038), the overall rate of minor hypoglycaemia remained low with both the BIAsp and the BHI treatments (13.1 vs. 8.3 episodes/patient year respectively). Overall safety and patient satisfaction were similar with the two insulin therapies. CONCLUSIONS: This trial confirmed that a thrice-daily BIAsp regimen can safely be used to intensify treatment for patients inadequately controlled on twice-daily BHI. A treat-to-target trial is required to explore the full potential of the BIAsp regimens and evaluate their use as a viable alternative to intensification with a basal-bolus regimen.

Inhibition of gamma-secretases alters both proliferation and differentiation of mesenchymal stem cells.

INTRODUCTION: Human mesenchymal stem cell (hMSC) proliferation and development is regulated by many signalling pathways. gamma-Secretases play an important role in Notch signalling as well as other processes that are involved in developmental decisions, but their role in hMSC proliferation and cell fate decisions has not been explored. OBJECTIVE: To investigate the role of gamma-secretases in hMSC proliferation and differentiation. MATERIALS AND METHODS: Using the gamma-secretase inhibitor N-[N-(3,5-Difluorophenacetyl-L-alanyl]-S-phenylglycine t-butyl ester (DAPT), we investigated their role in hMSC growth and differentiation to chondrogenic, osteogenic and adipogenic fates. RESULTS: We found that inhibiting gamma-secretases reduced the rate of hMSC proliferation, and altered hMSC differentiation in vitro. Addition of DAPT had an inhibitory effect on chondrogenesis resulting in impaired cartilage matrix production and altered chondrocyte morphology. DAPT treated chrodrocytic pellets had reduced levels of Hes1 and Hey1 suggesting that these effects are mediated via Notch signalling. Addition of the DAPT inhibitor to osteogenic cultures did not alter the appearance of bone markers, however, adipogenesis occurred in these cultures in a DAPT concentration-dependent manner. DAPT did not enhance adipogenesis in the presence of a potent adipogenic cocktail, but had an adipogenic effect when combined with dexamethasone only. CONCLUSION: We conclude that gamma-secretases play an important role in both hMSC proliferation and differentiation.

Vitamin D supply to the rat fetus and neonate.

The prevention of neonatal rickets by oral supplementation with vitamin D2 (ergocalciferol) has tended to obscure our ignorance of the natural mechanism by which young mammals receive an adequate supply of vitamin D. To investigate the possibility of specific intrauterine transfer and storage of vitamin D in fetal tissues, vitamin D-deficient female rats were given depot injections of 3H- or 14C-labeled vitamin D3 (cholecalciferol) before mating and the 3H-labeled animals were killed at stages during the last third of gestation. Analysis of lipid extracts from whole fetuses revealed a linear increase in the concentration of 25-hydroxyvitamin D3, 24,25-dihydroxyvitamin D3, and D3 itself between days 14 and 19 of gestation. During this period the elimination half-time of 3H-labeled molecules in maternal plasma fell from 27.1 to 4.4 d, suggesting that a specific mechanism was transferring vitamin D molecules into the fetuses. The vitamin was stored predominantly as 25-hydroxyvitamin D3 and 24,25-dihydroxyvitamin D3, with the highest concentrations in fetal muscle. Immediately after birth, pups from 3H- and 14C-labeled mothers were exchanged and later killed after 1-3 wk of suckling. Analysis of total lipid extracts for 3H and 14C content determined the relative contributions of vitamin D supplied before birth via the placenta and after birth in the maternal milk. The vitamin D content of the rat milk was relatively high, between 1.0 and 3.5 micrograms/liter. Nevertheless, the supply of vitamin D in utero, rather than from milk, was the main determinant of vitamin D status in early neonatal life. This is the first indication in a mammal of a specific transfer mechanism that allows the fetus to accumulate vitamin D from the mother during the last third of gestation.

Putative human liver progenitor cells in explanted liver.

BACKGROUND/AIMS: Hepatocyte progenitors have frequently been cultured from rodents but reports from human liver are rare. METHODS: Non-parenchymal cell fraction isolated from 19 explant livers (removed at orthotopic liver transplantation for acute or chronic liver disease) and histologically normal human liver was cultured. RESULTS: Proliferating epithelioid colonies were identifiable after 2-3 weeks culture as a very rare event (<1 per million cells plated) expressing mRNAs and protein antigens of mixed hepatocytic/biliary phenotype. Colony survival could be prolonged by transduction of the catalytic sub-unit of telomerase. Hepatocyte growth factor, epidermal growth factor and oncostatin M did not further enhance hepatocytic differentiation. The expression of markers associated with hepatocyte precursor status was investigated by flow cytometry. Cells expressing the stem cell-associated markers CD133 and CD117 were identified at low frequency. The proportion of cells expressing the integrin CD49f was higher in diseased liver than in normal liver, but the proportion expressing the hepatocyte growth factor receptor c-met was lower. Successful enrichment of plated populations for progenitors was not achieved. CONCLUSION: Although there is clear histological evidence of hepatocyte precursors in human explant livers, predictable culture of such cells with differentiation toward mature hepatocyte phenotype remains elusive.

Prostate cancer screening in men aged 50 to 69 years (STHLM3): A prospective population-based diagnostic study. Grönberg H, Adolfsson J, Aly M, Nordström T, Wiklund P, Brandberg Y, Thompson J, Wiklund F, Lindberg J, Clements M, Egevad L, Eklund M.Lancet Oncol. 2015 Dec;16(16):1667-76. [Epub 2015 Nov 10]. doi: 10.1016/S1470-2045(15)00361-7.

BACKGROUND: The prostate-specific antigen (PSA) test is used to screen for prostate cancer but has a high false-positive rate that translates into unnecessary prostate biopsies and overdiagnosis of low-risk prostate cancers. We aimed to develop and validate a model to identify high-risk prostate cancer (with a Gleason score of at least 7) with better test characteristics than that provided by PSA screening alone. METHODS: The Stockholm 3 (STHLM3) study is a prospective, population-based, paired, screen-positive, diagnostic study of men without prostate cancer aged 50 to 69 years randomly invited by date of birth from the Swedish Population Register kept by the Swedish Tax Agency. Men with prostate cancer at enrolment were excluded from the study. The predefined STHLM3 model (a combination of plasma protein biomarkers [PSA, free PSA, intact PSA, hK2, MSMB, MIC1], genetic polymorphisms [232 SNPs], and clinical variables [age, family, history, previous prostate biopsy, prostate exam]), and PSA concentration were both tested in all participants enrolled. The primary aim was to increase the specificity compared with PSA without decreasing the sensitivity to diagnose high-risk prostate cancer. The primary outcomes were number of detected high-risk cancers (sensitivity) and the number of performed prostate biopsies (specificity). The STHLM3 training cohort was used to train the STHLM3 model, which was prospectively tested in the STHLM3 validation cohort. Logistic regression was used to test for associations between biomarkers and clinical variables and prostate cancer with a Gleason score of at least 7. This study is registered with ISCRTN.com, number ISRCTN84445406. FINDINGS: The STHLM3 model performed significantly better than PSA alone for detection of cancers with a Gleason score of at least 7 (P<0.0001), the area under the curve was 0·56 (95% CI: 0·55-0·60) with PSA alone and 0·74 (95% CI: 0·72-0·75) with the STHLM3 model. All variables used in the STHLM3 model were significantly associated with prostate cancers with a Gleason score of at least 7 (P<0·05) in a multiple logistic regression model. At the same level of sensitivity as the PSA test using a cutoff of≥3ng/ml to diagnose high-risk prostate cancer, use of the STHLM3 model could reduce the number of biopsies by 32% (95% CI: 24-39) and could avoid 44% (35-54) of benign biopsies. INTERPRETATION: The STHLM3 model could reduce unnecessary biopsies without compromising the ability to diagnose prostate cancer with a Gleason score of at least 7, and could be a step towards personalised risk-based prostate cancer diagnostic programmes. FUNDING: Stockholm County Council (Stockholms Läns Landsting).

Quantitation of serum prostate-specific membrane antigen by a novel protein biochip immunoassay discriminates benign from malignant prostate disease.

The lack of a sensitive immunoassay for quantitating serum prostate-specific membrane antigen (PSMA) hinders its clinical utility as a diagnostic/prognostic biomarker. An innovative protein biochip immunoassay was used to quantitate and compare serum PSMA levels in healthy men and patients with either benign or malignant prostate disease. PSMA was captured from serum by anti-PSMA antibody bound to ProteinChip arrays, the captured PSMA detected by surface-enhanced laser desorption/ionization mass spectrometry, and quantitated by comparing the mass signal integrals to a standard curve established using purified recombinant PSMA. The average serum PSMA value for prostate cancer (623.1 ng/ml) was significantly different (P < 0.001) from that for benign prostate hyperplasia (117.1 ng/ml) and the normal groups (age <50, 272.9 ng/ml; age >50, 359.4 ng/ml). These initial results suggest that serum PSMA may be a more effective biomarker than prostate-specific antigen for differentiating benign from malignant prostate disease and warrants additional evaluation of the surface-enhanced laser desorption/ionization PSMA immunoassay to determine its diagnostic utility.