Insulin resistance and intestinal integrity in children with and without HIV infection in Uganda.

OBJECTIVES: The risk of cardiometabolic complications in children with perinatally acquired HIV infection (PHIVs) and in perinatally HIV-exposed but uninfected children (HEUs) and its relationship to systemic inflammation and markers of gut integrity are not well established. In this current study, we assed insulin resitance in PHIV compared to HEUs and HIV unexposed uninfected children and explored potential association with intestinal damage biomarkers. METHODS: This was a cross-sectional study in PHIVs, HEUs and HIV-unexposed, uninfected children (HUUs) aged 2-10 years enrolled in Uganda. PHIVs were on stable antiretroviral therapy (ART) with HIV viral load  < 400 HIV-1 RNA copies/mL. Insulin resistance was estimated using the homeostasis model assessment of insulin resistance (HOMA-IR). We measured markers of systemic inflammation, monocyte activation and gut integrity. Kruskal-Wallis tests were used to compare markers by HIV status; Pearson correlation and multiple linear regressions were used to assess associations of the HOMA-IR index with biomarkers of intestinal damage and translocation. RESULTS: Overall, 172 participants were enrolled in the study (57 PHIVs, 59 HEUs and 56 HUUs). The median age was 7.8 [interquartile range (IQR) 6.39, 8.84] years, 55% were female and the median body mass index (BMI) was 15 (IQR 14.3, 15.8) kg/m2 . Among PHIVs, the median CD4% was 37%, and 93% had viral load ≤ 20 copies/mL. PHIVs had higher waist:hip ratio, high-density lipoprotein (HDL) cholesterol, triglycerides and HOMA-IR index than the other groups (P ≤ 0.02). Factors correlated with insulin resistance included higher BMI and HDL cholesterol and lower soluble tumour necrosis factor receptor I (sTNFRI) (P ≤ 0.02). There was no correlation between any of the other inflammatory or gut biomarkers and HOMA-IR index (P ≥ 0.05). After adjusting for age and sTNFRI, BMI remained independently associated with the HOMA-IR index (ß = 0.16; P < 0.01). CONCLUSIONS: Despite viral suppression, Ugandan PHIVs have disturbances in glucose metabolism. Higher BMI, and not immune activation or alteration of gut integrity, was associated with insulin resistance in this population.

Alcohol and dietary factors associate with gut integrity and inflammation in HIV-infected adults.

OBJECTIVES: HIV-infected adults have heightened monocyte activation and inflammation, at least partially as a consequence of altered gut integrity. The role of dietary factors in microbial translocation and inflammation and their downstream effect on markers of cardiovascular disease (CVD) have not been explored. Our purpose was to describe the longitudinal dietary patterns of HIV-infected adults, and to examine the relationship between dietary intake, gut integrity, inflammation and subclinical markers of CVD in HIV-infected adults. METHODS: We conducted a secondary analysis of 147 HIV-infected participants in a 96-week randomized clinical trial of rosuvastatin as primary CVD prevention. Dietary intake was assessed using dietary recall; plasma gut integrity, monocyte activation and inflammation markers were measured using an enzyme-linked immunosorbent assay (ELISA); and CVD risk was assessed using carotid ultrasound and the coronary artery calcium score. Linear mixed models were used to analyse longitudinally measured biomarkers. RESULTS: The median age was 45 years and 78% of patients were male. At baseline, participants consumed a mean (standard deviation) of 108 (70) g of fat daily, 19 (15.6) g of fibre, 266 (186) g of carbohydrates and 15.6 (5.9) g of protein; 45% of the sample consumed alcohol. Over time, alcohol consumption was associated with several markers of gut integrity and inflammation (all P < 0.05). CONCLUSIONS: HIV-infected adults in a contemporary, high-resource setting have poor dietary patterns. Alcohol use was associated with worse gut integrity and increased inflammation, while other aspects of diet (fibre, carbohydrates and fat) were not. These data add to growing evidence illustrating the need for a better understanding of the effect of lifestyle factors on comorbidities in HIV-infected adults.

Effect of cholesterol-loaded cyclodextrins on cryosurvival of dog spermatozoa.

Cryopreservation affects integrity of cholesterol and phospholipids in the plasma membrane of sperm leading to decreased fertility of frozen-thawed semen. Cholesterol-loaded cyclodextrins (CLC) have been shown to improve post-thaw semen quality in various species. The aim of this study was to investigate the optimal concentration of CLC for better post-thaw semen quality in dogs. Semen collection, through digital manipulation, was conducted once a week in four adult German shepherd dogs (n = 20 ejaculates; five ejaculates/dog). Semen samples with mass motility>3 (0: without movement; 5: fast progressive movement), motility >70% and concentration >200 × 106 /ml were pooled and processed in Tris-citrate extender containing 0, 1, 2 or 3 mg of CLC. The post-thaw quality was assessed on the basis of percentage motility, morphological abnormalities, live/dead ratio and plasma membrane, acrosome and DNA integrity, evaluated using anova and further analysed by Tukey's range test, if applicable. The addition of CLC showed an overall improvement in post-thaw semen quality. Among various treatment groups, and when compared to the control, the percentages of motile (55.5%), viable (65%), plasma membrane intact (56.7%), acrosome intact (49.2%) and DNA intact (98%) spermatozoa were significantly higher in 2 mg/ml CLC group (p < .05). It is concluded that incorporation of cholesterol in semen extender results in a beneficial increase in post-thaw semen quality in dogs.

A Probabilistic Approach to Mitigate Composition Attacks on Privacy in Non-Coordinated Environments.

Organizations share data about individuals to drive business and comply with law and regulation. However, an adversary may expose confidential information by tracking an individual across disparate data publications using quasi-identifying attributes (e.g., age, geocode and sex) associated with the records. Various studies have shown that well-established privacy protection models (e.g., k-anonymity and its extensions) fail to protect an individual's privacy against this "composition attack". This type of attack can be thwarted when organizations coordinate prior to data publication, but such a practice is not always feasible. In this paper, we introduce a probabilistic model called (d, α)-linkable, which mitigates composition attack without coordination. The model ensures that d confidential values are associated with a quasi-identifying group with a likelihood of α. We realize this model through an efficient extension to k-anonymization and use extensive experiments to show our strategy significantly reduces the likelihood of a successful composition attack and can preserve more utility than alternative privacy models, such as differential privacy.

Evaluation of Antimicrobial Efficacy and Clinical Outcomes of Triphala and 2.5% Sodium Hypochlorite as Intraradicular Irrigants in Pulpectomy of Primary Teeth.

A natural irrigation solution with a broad spectrum of antimicrobial coverage, triphala was selected for the pulpectomy procedure. Because of its natural ingredients, it is well-known for promoting tissue healing. It also supposedly has certain additional qualities as compared to usual irrigation solutions that are made chemically. Although 2.5% NaOCl is thought to be perfect since it meets most of the requirements for an irrigation solution but it cannot be optimized for pulpectomy procedure. Primary teeth that were recommended for pulpectomy underwent this randomized controlled experiment. Two groups of eighty-four primary teeth were randomly assigned to receive irrigations: triphala in Group A; 2.5% Sodium hypochlorite in Group B. Sample were taken from infected primary root canals. A sterile test tube with bhi broth as the transport media was used to collect pre- and post-irrigation samples using sterile absorbent paper tips. On agar media, microorganisms were cultivated and their mean colony count was assessed. Following the procedure, the patient's follow-up visits at one, two and three months were used to evaluate the clinical result. The post-microbial colony count was dramatically reduced (p<0.001) by both irrigation treatments. Triphala in Group A is demonstrating desirable efficacy. Clinical success was found satisfactory in both the groups studied (p<0.001). But statistically significant difference was not found (p=0.175). Considering undesirable properties of sodium hypochlorite triphala can be a better alternative as a root canal irrigants in pulpectomy of primary teeth.

Comparison of transvaginal and transabdominal ultrasonography in the diagnosis of ectopic pregnancy.

This cross sectional study was carried out in the department of Radiology and Imaging, Dhaka Medical College Hospital from July 2008 to June 2010 to compare the accuracy of transvaginal ultrasonography and transabdominal ultrasonography in the diagnosis of clinically suspected cases of ectopic pregnancy. Initially 60 patients with clinical suspicion of ectopic pregnancy were included in this study after analyzing selection criteria 30 patients underwent both transvaginal and transabdominal ultrasonography. 'Histopathological diagnosis' was considered gold standard against which accuracies of two diagnostic modalities were compared. Sensitivity, specificity, positive predictive value, negative predictive value and accuracy of transabdominal ultrasonography as a diagnostic modality in evaluation of suspected ectopic pregnancy were 73.1%, 75%, 95%, 30% and 73.3% respectively where as transvaginal ultrasonography was found to have 92.3% sensitivity, 75% specificity, 96% positive predictive value, 60% negative predictive value and 90% accuracy. Transvaginal ultrasonography was superior to transabdominal ultrasonography in the evaluation of suspected ectopic pregnancies so, transvaginal ultrasonography is important for early and accurate diagnosis of ectopic pregnancy.

Event studies in international finance research.

Event studies are widely used in finance research to investigate the implications of announcements of corporate initiatives, regulatory changes, or macroeconomic shocks on stock prices. These studies are often used in a single-country setting (usually the U.S.), but little work has yet been conducted in an international context, perhaps due to the complexities inherent in implementing cross-country studies. This paper explores the methodological challenges of conducting event studies in international finance research. We emphasize how scholars should choose an event, select the study period (short vs. long term), estimate abnormal returns, infer statistically whether the event under consideration produces a reliable price reaction, and explore the role of formal and informal institutions in explaining cross-country differences in price reactions. We also provide an extension of event studies to an important but less studied asset class in an international setting - the fixed-income market. We conclude by offering practical recommendations for researchers conducting cross-country finance event studies and identifying opportunities for future research. Given the increasing number of global events, such as the COVID-19 pandemic, Brexit, and the Paris and Trans-Pacific Partnership agreements, we believe our paper is especially timely.

Les études d'événements sont largement utilisées dans la recherche financière pour étudier les implications des annonces d'initiatives d'entreprises, de changements réglementaires ou de chocs macroéconomiques sur les prix des actions. Ces études sont souvent utilisées dans le cadre d'un seul pays (généralement les États-Unis), mais peu de travaux ont encore été menés dans un contexte international, probablement en raison des complexités inhérentes à la mise en oeuvre d'études transnationales. Cet article explore les défis méthodologiques de la conduite d'études d'événements dans la recherche en finance internationale. Nous mettons l'accent sur la manière dont les chercheurs doivent choisir un événement, sélectionner la période d'étude (court ou long terme), estimer les rendements anormaux, déduire statistiquement si l'événement considéré produit une réaction fiable des prix, et explorer le rôle des institutions formelles et informelles dans l'explication des réactions des prix à travers les réactions des prix. Nous proposons également une extension des études d'événements à une classe d'actifs importante mais moins étudiée dans un contexte international - le marché des titres à revenu fixe. Nous concluons en offrant des recommandations pratiques aux chercheurs qui mènent des études d'événements financiers transnationaux et en identifiant les possibilités de recherches futures. Compte tenu du nombre croissant d'événements mondiaux, tels que la pandémie de COVID-19, le Brexit et les accords de Paris et de partenariat transpacifique, nous pensons que notre article est particulièrement opportun.

Los estudios de eventos son ampliamente usados en la investigación financiera para indagar las implicaciones de los anuncios de iniciativas empresariales, cambios regulatorios o choques macroeconómicos en los precios de las acciones. Estos estudios se utilizan a menudo en el marco de un solo país (normalmente EE.UU.), pero todavía se han realizado pocos trabajos en un contexto internacional, quizá debido a las complejidades inherentes a la realización de estudios transfronterizos. Este artículo explora los retos metodológicos de llevar a cabo estudios de eventos en la investigación financiera internacional. Enfatizamos cómo los académicos deben elegir un evento, seleccionar el período de estudio (corto frente a largo plazo), estimar los rendimientos anormales, inferir estadísticamente si el evento en cuestión produce una reacción fiable de los precios, y explorar el papel de las instituciones formales e informales para explicar las diferencias entre países en las reacciones de los precios. También ofrecemos una extensión de los estudios de eventos a una clase de activos importante pero menos estudiada en un entorno internacional: el mercado de renta fija. Concluimos ofreciendo recomendaciones prácticas para los investigadores que realizan estudios de eventos financieros entre países e identificando oportunidades para futuras investigaciones. Dado el creciente número de eventos mundiales, como la pandemia del COVID-19, el Brexit y el Acuerdo de París y el Acuerdo de Asociación Transpacífico, creemos que nuestro artículo es especialmente oportuno.

Estudos de eventos são amplamente utilizados em pesquisas sobre finanças para investigar as implicações de anúncios de iniciativas corporativas, mudanças regulatórias ou choques macroeconômicos nos preços das ações. Esses estudos são frequentemente usados em um único país (geralmente os EUA), mas pouco trabalho foi realizado em um contexto internacional, talvez devido às complexidades inerentes à implementação de estudos entre países. Este artigo explora os desafios metodológicos na condução de estudos de eventos na pesquisa de finanças internacionais. Enfatizamos como acadêmicos devem escolher um evento, selecionar o período de estudo (curto prazo versus longo prazo), estimar retornos anormais, inferir estatisticamente se o evento em análise produz uma reação de preço confiável e explorar o papel de instituições formais e informais na explicação de diferenças entre países nas reações de preços. Também fornecemos uma extensão dos estudos de eventos para uma classe de ativos importante, mas menos estudada em um cenário internacional ­ o mercado de renda fixa. Concluímos oferecendo recomendações práticas para pesquisadores que realizam estudos de eventos em finanças entre países e identificando oportunidades para pesquisas futuras. Dado o número crescente de eventos globais, como a pandemia de COVID-19, Brexit e os acordos de Paris e parceria Trans-Pacific, acreditamos que nosso artigo é especialmente oportuno.

Morphological and molecular evidences of Ascaridia galli in migratory quail Coturnix coturnix japonica from Baluchistan Pakistan.

The current study was conducted to examine the point prevalence of gastrointestinal parasites of migratory quails. Due to its economic importance, the control of ascaridiosis is critical. Migration of birds is considered to enhance the global spread and cross-species transmission of pathogens. The current study was aimed to detect A.galli in migratory quails, a potential contributory risk factor for transmission of this parasite to local birds. A total of 230 migratory quails were trapped using nets from migratory routes in Balochistan and examined under the compound microscope for the presence of A. galli. Conventionally, A. galli was identified by its morphology with the presence of three large lips and absence of posterior esophageal bulb. Results revealed that out of 230, 120 (52.17%) quails were positive for A. galli by targeting COX1 gene (533 bp) by using conventional PCR. Further, the amplicon was sequenced which showed 99% similarity with A. galli publically available in NCBI Gen Bank. Phylogenetic analysis of sequences of our isolated parasite indicated the close relationship with A.galli isolated from chickens. In conclusion migratory quails and other migratory birds may play a key role in spreading and transmission of these parasites and other pathogens to domestic chicken. Therefore, strict biosecurity measures should be adopted especially for commercial poultry farms.

The influence of gamete co-incubation length on the in vitro fertility and sex ratio of bovine bulls with different penetration speed.

The objectives of this work were to evaluate whether the sperm penetration speed is correlated to the in vitro fertility and whether adapting the gamete co-incubation length to the kinetics of the bull improves in vitro fertility and affects the sex ratio. In vitro matured oocytes were co-incubated with spermatozoa from four different bulls (A-D). At various post-insemination (p.i.) times (4, 8, 12, 16 and 20 h), samples of oocytes were fixed and stained with DAPI for nuclei examination, while the remaining ones were transferred into culture to evaluate embryo development. The blastocysts produced were sexed by PCR. Two bulls (A and B) had faster kinetics than the others (C and D), as shown by the higher penetration rates recorded at 4 h p.i. (43%, 30%, 11% and 6%, respectively for bulls A, B, C and D; p<0.01). The differences in the kinetics among bulls did not reflect their in vitro fertility. The incidence of polyspermy was higher for faster penetrating bulls (36%, 24%, 16% and 4%, respectively for bulls A, B, C and D; p<0.01) and at longer co-incubation times (0%, 16%, 19%, 30% and 34%, respectively at 4, 8, 12, 16 and 20 h p.i.; p<0.01). The fertilizing ability of individual bulls may be improved by adapting the co-incubation length to their penetration speed. A sperm-oocyte co-incubation length of 8 h ensured the greatest blastocyst yields for the two faster penetrating bulls. On the contrary, 16 h co-incubation was required to increase (p<0.01) cleavage rate of the two slower bulls. Bulls with a faster kinetics did not alter the embryo sex ratio towards males. The female/male (F/M) ratios recorded were 2.1, 1.4, 1.2, 1.3 and 1.6, respectively at 4, 8, 12, 16 and 20 h p.i.

Workplace mistreatment and mental health in female surgeons in Pakistan.

BACKGROUND: Despite workplace mistreatment, which includes harassment, bullying and gender discrimination(GD)/bias, being serious problems for female surgeons, there are limited data from lower-middle-income countries like Pakistan. This study explored harassment and GD/bias experienced by female surgeons in Pakistan, and the effects of these experiences on mental health and well-being. METHODS: A nationwide survey was conducted between July and September 2019 in collaboration with the Association of Women Surgeons of Pakistan, an organization consisting of female surgeons and trainees in Pakistan. An anonymous online survey was emailed directly, disseminated via social media platforms (such as Facebook, Twitter and Instagram), and sent to surgical programmes in Pakistan. RESULTS: A total of 146 women surgeons responded to the survey; 67.1 per cent were trainees and the rest attending surgeons. Overall, 57.5 per cent of surgeons reported experiencing harassment, most common being verbal (64.0 per cent) and mental (45.9 per cent), but this mostly went unreported (91.5 per cent). On multivariable analysis adjusted for age and specialty, workplace harassment (odds ratio 2.02 (95 per cent c.i. 1.09 to 4.45)) and bullying (odds ratio 5.14 (95 per cent c.i. 2.00-13.17)) were significantly associated with severe self-perceived burnout, while having a support system was protective against feelings of depression (odds ratio 0.35 (95 per cent c.i. 0.16 to 0.74)). The overwhelming majority (91.3 per cent) believed that more institutional support groups were needed to help surgeons with stress reduction (78.8 per cent), receiving mentorship (74.7 per cent) and work-life balance (67.8 per cent). CONCLUSION: Workplace mistreatment, in particular harassment and bullying, has a damaging impact on the mental well-being of female surgeons, particularly trainees. The absence of support groups in Pakistan should be urgently addressed so that surgeons, especially trainees, may cope better with potentially harmful workplace stressors.

A massive rock and ice avalanche caused the 2021 disaster at Chamoli, Indian Himalaya.

On 7 February 2021, a catastrophic mass flow descended the Ronti Gad, Rishiganga, and Dhauliganga valleys in Chamoli, Uttarakhand, India, causing widespread devastation and severely damaging two hydropower projects. More than 200 people were killed or are missing. Our analysis of satellite imagery, seismic records, numerical model results, and eyewitness videos reveals that ~27 × 106 cubic meters of rock and glacier ice collapsed from the steep north face of Ronti Peak. The rock and ice avalanche rapidly transformed into an extraordinarily large and mobile debris flow that transported boulders greater than 20 meters in diameter and scoured the valley walls up to 220 meters above the valley floor. The intersection of the hazard cascade with downvalley infrastructure resulted in a disaster, which highlights key questions about adequate monitoring and sustainable development in the Himalaya as well as other remote, high-mountain environments.

Determination of concentrations of Fe, Mg, and Zn in some ferrite samples using neutron activation analysis and X-ray fluorescence techniques.

Mg-Zn ferrite is considered as one of the important materials with potential uses in many applications. In this work, samples of ferrite Mg(1-x)ZnxFe2O4 (where x=0.0, 0.2, 0.4, 0.6, 0.8 and 1) were synthesized by the sol-gel method for use in some hyperthermia applications. The composition and purity of the prepared samples hardly affected their properties. Therefore, the elemental concentration of these samples was measured by the X-ray fluorescence technique and thermal neutron activation analysis to check the quality of the prepared samples. The results of both methods were compared with each other and with the molecular ratios of the as-prepared samples. In addition, no existing elemental impurity, with considerable concentration, was measured.

Weibel-Palade bodies as a marker for neovascularization induced by tumor and rheumatoid angiogenesis factors.

An analysis was made of ultrastructural changes in capillary endothelial cells in experimentally induced angiogenesis and in a human pathological situation known to involve increased angiogenesis. Chick chorioalloantoic membrane (CAM) showing a positive angiogenic response to low-molecular weight angiogenesis factor isolated from rat Walker sarcoma or from human rheumatoid joint was compared with untreated CAM. Serotonin-treated CAM provided an additional control in that serotonin has the capacity to stimulate endothelial cell growth in vitro but did not induce angiogenesis on the CAM. Human rheumatoid joints were studied using normal healthy human joints as controls. The number of Weibel-Palade (W-P) bodies per unit of cytoplasmic area were higher in tumor angiogenesis factor-treated CAMs (not significant) and rheumatoid angiogenesis factor-treated CAMs (P less than 0.008) than in untreated controls. These differences were more pronounced if W-P body volumetric density was analyzed (P in both cases less than 0.008). Serotonin-treated control CAMs did not show higher numbers of W-P body or greater WPV than untreated controls. Numbers of W-P body and W-P body volumetric density were higher (P less than 0.008) in rheumatoid joints than normal joints. Median values for W-P body number were 16-fold higher and, for W-P body volumetric density, they were up to 30-fold higher in rheumatoid joints.

Comparative efficacy of G6G and Ovsynch protocols on synchronization and pregnancy rate in Nili-Ravi buffalo.

The aim of this work was to evaluate the efficacy of G6G and Ovsynch protocols on synchronization and pregnancy rate in Nili-Ravi buffalo. For this, a total of fifty buffalo were assigned into two treatment groups: (1) G6G (n=25) and (2) Ovsynch (n=25). The blood samples were collected from all the treated buffalo on day (D) 7 (PGF2α injection of Ovsynch) to measure the progesterone (P4). All the buffalo were fixed time inseminated (FTAI) following last GnRH of Ovsynch. Ultrasonography was performed for follicular, corpus luteum (CL) dynamics and pregnancy diagnosis. The results revealed that synchronization rate and response to final PGF2α were significantly (P<0.05) higher in G6G compared to Ovsynch. The ovulation to first GnRH of Ovsynch was higher (P<0.05) in G6G (84%) than Ovsynch (56%). The mean dominant follicle size following last GnRH of Ovsynch showed significantly larger diameter in G6G (11.7±0.2mm) compared to Ovsynch (10.9±0.2mm). Moreover, the higher (P<0.05) P4 concentration was observed in G6G than Ovsynch. The presence of double CL on D 7 was higher (P<0.05) in G6G (64%) than Ovsynch (24%) group. The pregnancy rate (P=0.08) tended to be greater in G6G compared to Ovsynch. In conclusion, G6G treatment showed better ovulatory and luteolytic response to first GnRH and PGF2α compared to Ovsynch treatment. Moreover, the addition of GnRH and PGF2α prior to Ovsynch is helpful to improve synchronization application of Ovsynch protocol in Nili-Ravi buffalo.

Application of angiogenic oligosaccharides of hyaluronan increases blood vessel numbers in rat skin.

Angiogenic oligosaccharides of hyaluronan were applied to the backs of young, adult male rats and the number of blood vessels, within a depth of 136 microns beneath the base of the epidermis, were evaluated. Application of hyaluronan oligosaccharides significantly increased the mean number of blood vessels/mm skin length in six of 11 treated rats when compared with controls. Application of radiolabeled hyaluronan oligosaccharides to skin of one rat demonstrated a penetration to a depth of approximately 800 microns, suggesting that the blood vessels beneath the epidermis would be exposed to the hyaluronan. Hyaluronan has previously been shown to stimulate endothelial cell proliferation; we demonstrate here that these hyaluronan oligosaccharides also specifically stimulate endothelial cell migration. This action of hyaluronan oligosaccharides may prove useful in retarding blood vessel paucity and degeneration observed during the ageing process and following radiotherapy.

Does hyaluronan have a role in endothelial cell proliferation of the synovium?

Hyaluranate (HA) is a major constituent of synovial fluid, but its concentration and molecular size differ in normal and inflamed joints. HA can induce or inhibit angiogenesis depending on both its size and its concentration. Endothelial-cell endocytose-labeled macromolecular HA and HA oligosaccharides and binding studies have identified an HA-specific receptor on the endothelial cell surface (KD, 10(-10) mol/L; approximately 2,000/cell). The molecular weight of HA-binding proteins was found to be 90 to 125, 78, and 46 kd.

Studies on prolyl endopeptidase from shakashimeji (Lyophyllum cinerascens): purification and enzymatic properties.

High prolyl endopeptidase (post-proline cleaving enzyme) [EC 3.4.21.26] activity was detected in fruit bodies of shakashimeji (Lyophyllum cinerascens), tsukuritake (mushroom: Agaricus bisporus), hirohachichitake (Lactarius hygrophoroides), and yaburebenitake (Russula lepida) which belong to the genus Basidiomycetes. Cell-free extract of shakashimeji showed high activities of proline iminopeptidase and arylamidase as well as prolyl endopeptidase. The prolyl endopeptidase was purified from the extract of shakashimeji by sequential chromatographies on DEAE-Toyopearl, DEAE-Sephadex and hydroxyapatite, and high-performance liquid chromatography with a DEAE-5PW column. The purified enzyme was homogeneous as judged by disc gel electrophoresis. The enzyme was most active at pH 6.8 as checked with Z-Gly-Pro-beta-naphthylamide as a substrate and was stable in the range of pH 5.8-7.4. The isoelectric point of the enzyme was 5.2 and the molecular weight was estimated to be 76,000 by gel filtration on Sephadex G-150 and by sodium dodecyl sulfate (SDS) gel electrophoresis, suggesting that the enzyme was a monomer. The enzyme was completely inhibited by diisopropyl fluorophosphate (DFP), Z-Gly-Pro-CH2Cl, and Z-Pro-prolinal, while it was not inhibited by p-chloromercuribenzoate (PCMB), phenylmethylsulfonyl fluoride (PMSF), or metal chelators. It was estimated that at least five subsites were concerned with the enzyme-substrate binding. Among them, the S1, S2, and S1' sites showed high stereospecificity, as in mammalian, microbial, and plant enzymes. The enzyme hydrolyzed TRH at the carboxyl side of the proline residue. The mushroom enzyme, that was sensitive to DFP, Z-Pro-prolinal, and Z-Gly-Pro-CH2Cl, but not to PCMB, were quite similar in characteristics to the Flavobacterium enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

Purification and characterization of an extracellular prolyl endopeptidase from Agaricus bisporus.

Prolyl endopeptidase [EC 3.4.21.26] was purified to homogeneity from the culture filtrate of Agaricus bisporus by a procedure that comprised ammonium sulfate fractionation, anion-exchange chromatographies on DEAE-Toyopearl and DEAE-Sephadex, hydroxylapatite chromatography, and high-performance liquid chromatography (HPLC) on a TSKgel G 2000 SW column. The overall activity recovery was 8.6%. The enzyme was most active at or around pH 7.5 and was stable in the range of pH 5-9 when checked with Z-Gly-Pro-beta-naphthylamide as a substrate. The isoelectric point of the enzyme was about 4.8. The enzyme was a monomeric protein of molecular weight 78,000 +/- 2,000 as judged by gel permeation chromatography on Sephadex G-150 and electrophoresis on sodium dodecyl sulfate (SDS) polyacrylamide gel. The enzyme hydrolyzed Pro-X bonds and at least five subsites (S3, S2, S1, S1', and S2') were found to be involved in enzyme-substrate binding. Among them, S2, S1, and S1' subsites of the enzyme showed high stereospecificity. The enzyme was strongly inhibited by diisopropylfluorophosphate (DFP), Z-Gly-Pro-CH2Cl, Z-Pro-prolinal, Z-Pro-pyrrolidine, Z-Thiopro-pyrrolidine, Z-Pro-thiazolidine, Z-Thioprothiazolidine, and p-chloromercuribenzoate (PCMB), while it was not inhibited by phenyl-methylsulfonyl fluoride (PMSF), E-64, iodoacetamide, or metal chelators. Although the A. bisporus enzyme showed no immunological cross reaction with anti-bovine prolyl endopeptidase antiserum, the other characteristics were quite similar to those of mammalian and plant enzymes.

Is one single prostate biopsy helpful for choosing a medical treatment of benign prostatic hyperplasia? A quantitative computerized morphometric study.

OBJECTIVES: Individual differences in proportion of stroma, epithelium, and luminal space components prostatic hyperplasia (BPH) may explain the differences in clinical outcome of the patients treated with alpha-reductase inhibitors or alpha-blocking agents and other alternative treatments. Knowledge of the individual proportions of these elements may orient the clinician toward different therapeutic approaches. To determine whether a single prostate biopsy is representative of the whole adenoma, using quantitative morphometry, we have compared the percentage of smooth muscle cells and glandular cells in one prostate needle biopsy and in the corresponding prostate adenoma removed by open surgery. METHODS: Quantification was made using a computerized image analysis system and immunohistochemical staining (actin antiactin for the smooth muscle cells and anti-prostate-specific antigen (PSA) for the epithelial cells) in 14 patients who underwent retropubic prostatectomy. RESULTS: Mean percentage of smooth muscle fibers, glandular epithelium, and glandular lumina in the prostate biopsy were, respectively, 34% (range, 20% to 42%, SD 5.9), 29% (range, 13% to 42%, SD 7.2), and 20% (range, 13% to 30%, SD 4.8). In the corresponding prostate adenoma, they were, respectively, 38% (range, 28% to 45%, SD 4.7), 32% (range, 25% to 40%, SD 4.5), and 19% (range, 13% to 34%, SD 6.1). The mean percentages of epithelial or glandular cells in the prostate biopsy and the corresponding adenoma were not statistically different (P > 0.05), whereas those for the smooth muscle cell percentage were different (P = 0.02). However, a statistically significant correlation between the whole adenoma and the needle biopsy sample was found in the percentage of smooth muscle cells (P = 0.028). Analyzing the morphometric data in conjunction with the serum PSA level and the volume of the adenoma, we found a statistically significant positive correlation between the volume of the adenoma and the ratio (percentage of epithelial cells/serum PSA level) (P = 0.009, r = 0.67). CONCLUSIONS: Major differences are found in the primary tissue composition of different hyperplastic prostates. Computerized histologic quantification of the different components of BPH in the entire adenoma and a needle biopsy demonstrate that a single prostate biopsy is representative of the entire prostate adenoma. Morphometric data from a single biopsy of the prostate adenoma in combination with the serum PSA level and the volume of the prostate adenoma could therefore help to orient medical treatment of BPH by patient selection based on the knowledge of the distribution of the various components of BPH.

Prostate capsule: computerized morphometric analysis of its components.

OBJECTIVES: There is little objective information on what is designated as being the prostate capsule. We have quantified objectively with computerized image analysis the percentage of smooth muscle fibers (SMFs) surrounding the surgical specimens of radical prostatectomies. METHODS: Ten radical prostatectomy specimens were used. Six different parts from prostate lobe were analyzed. Thirty different fields were examined from each tissue section. For tissue sections, different stains were used: hematoxylin and eosin, Masson's trichrome, and immunohistochemical staining with antiactin for SMFs. RESULTS: The mean thickness of the prostate capsule was 0.5 to 2 mm. The mean percentages of SMFs in the different regions of the prostate capsule was 30.6%. No difference was noted between the percentage of SMFs in the prostatic capsule compared with that in the prostate parenchyma itself (30.7%). CONCLUSIONS: This study provides objective evidence to demonstrate that the prostate does not have a true capsule. This capsule represents a fibromuscular band that probably should be considered as the extension of the internal parenchyma.