Detection of low numbers of bacterial cells in a pharmaceutical drug product using Raman spectroscopy and PLS-DA multivariate analysis.

Sterility testing is a laborious and slow process to detect contaminants present in drug products. Raman spectroscopy is a promising label-free tool to detect microorganisms and thus gaining relevance as a future alternative culture-free method for sterility testing in the pharmaceutical industry. However, reaching detection limits similar to standard procedures while keeping a high accuracy remains challenging, due to weak bacterial Raman signals. In this work, we show a new non-invasive approach focusing on detection of different bacteria in concentrations below 100 CFU per ml within drug product containers using Raman spectroscopy and multivariate data analysis. Even though Raman spectra from drug product with and without bacteria are similar, a partial least squared discriminant analysis (PLS-DA) model shows great performance to distinguish samples with bacterial contaminants in concentrations down to 10 CFU per ml. We used spiked samples with bacterial spores for model validation achieving a detection accuracy of 99%. Our results indicate the great potential of this rapid, and cost-effective approach to be used in quality control in the pharmaceutical industry.

The Pandora multi-algorithm approach to automated pattern recognition of cosmic-ray muon and neutrino events in the MicroBooNE detector.

The development and operation of liquid-argon time-projection chambers for neutrino physics has created a need for new approaches to pattern recognition in order to fully exploit the imaging capabilities offered by this technology. Whereas the human brain can excel at identifying features in the recorded events, it is a significant challenge to develop an automated, algorithmic solution. The Pandora Software Development Kit provides functionality to aid the design and implementation of pattern-recognition algorithms. It promotes the use of a multi-algorithm approach to pattern recognition, in which individual algorithms each address a specific task in a particular topology. Many tens of algorithms then carefully build up a picture of the event and, together, provide a robust automated pattern-recognition solution. This paper describes details of the chain of over one hundred Pandora algorithms and tools used to reconstruct cosmic-ray muon and neutrino events in the MicroBooNE detector. Metrics that assess the current pattern-recognition performance are presented for simulated MicroBooNE events, using a selection of final-state event topologies.

Determination of the elasticity parameters of brain tissue with combined simulation and registration.

Reliable elasticity parameters describing the behavior of a given material are an important issue in the context of physically-based simulation. In this paper we introduce a method for the determination of the mechanical properties of brain tissue. Elasticity parameters Young's modulus E and Poisson's ratio nu are estimated in an iterative framework coupling a finite element simulation with image registration. Within this framework, the outcome of the simulation is parameterized with both elasticity moduli that are automatically varied until optimal image correspondence between the simulated and the intraoperative data is achieved. We calculated optimal mechanical properties of brain tissue in six cases. The statistical analysis of the obtained values showed a good correlation of the results, thus proving the value of the method. An approach combining simulation and registration for the determination of the mechanical brain tissue properties is presented. This contributes to performing reliable physically-based simulation of soft tissue movement.

Simultaneous lightness contrast with double increments.

In this paper we demonstrate the existence of simultaneous lightness contrast in displays in which the target patches are both more luminant than their surrounds. These effects are not predicted by theories of lightness that assume that the highest luminance in a scene is perceived as white, and anchors all the other luminances. We show that the strength of double-increment illusions depends crucially on the luminance of both the surrounds and the target patches. Such luminance prerequisites were not met in previous studies, which explains why simultaneous contrast with incremental targets has so far been regarded as extremely weak or nonexistent.

A comparison of stavudine plus lamivudine versus zidovudine plus lamivudine in combination with indinavir in antiretroviral naive individuals with HIV infection: selection of thymidine analog regimen therapy (START I).

BACKGROUND: No clinical trial results directly comparing two nucleoside analog pairs in a drug regimen for HIV that includes a protease inhibitor are available. OBJECTIVE: To compare the safety and efficacy of stavudine (d4T) + lamivudine (3TC) with zidovudine (ZDV) + 3TC, each in combination with indinavir (IDV). DESIGN: Randomized, open-label, multi-center. SETTING: Fifteen HIV clinical research centers. PATIENTS: Two-hundred and four antiretroviral-naive HIV-1-infected-patients with CD4 cell counts > or = 200 x 10(6)/l and HIV-1 RNA > or = 10,000 copies/ml (bDNA assay), modified to 5000 copies/ml. INTERVENTION: d4T 40 mg twice a day, 3TC 150 mg twice a day plus IDV 800 mg every 8 h compared with ZDV 200 mg every 8 h (modified to 300 mg every 12 h) plus 3TC and IDV. MEASUREMENTS: Primary endpoint: plasma HIV-1 RNA < 500 copies/ml. Additional endpoints: HIV-1 RNA < or = 50 copies/ml; change from baseline in HIV-1 RNA and CD4 cell counts; safety and adverse events. RESULTS: For HIV-1 RNA, 62% of patients on d4T + 3TC + IDV and 54% of patients on ZDV + 3TC + IDV had < 500 copies/ml HIV RNA at weeks 40 through 48 [90% confidence interval, -0.204 to 0.036; P= 0.213], with 49% and 47% respectively achieving < or = 50 copies/ml at 48 weeks (90% CI, -0.134 to 0.096; P = 0.834). Median change in CD4 cell counts at 48 weeks was +227 x 10(6)/l and +198 x 10(6)/l for the d4T- and ZDV-containing arms, respectively. The median time-weighted average change from baseline in CD4 cell counts was significantly greater at 48 weeks in the d4T-containing arm (142 x 10(6)/l versus 110 x 10(6)/l; P = 0.033). Serious adverse events were not significantly different between treatment arms, but there were significant differences for frequency of adverse events of all severity with increased nausea and vomiting in the ZDV-containing arm, and increased diarrhea and rash in the d4T-containing arm. CONCLUSIONS: These results support the choice of d4T + 3TC as a nucleoside analog pair in combination with a protease inhibitor in an initial HIV treatment regimen.

A comparison of stavudine, didanosine and indinavir with zidovudine, lamivudine and indinavir for the initial treatment of HIV-1 infected individuals: selection of thymidine analog regimen therapy (START II).

OBJECTIVE: Comparison of stavudine (d4T), didanosine (ddI) and indinavir (IDV) with zidovudine (ZDV), lamivudine (3TC) and IDV in HIV-1 infected patients. DESIGN: Randomized, open-label. SETTING: Fourteen HIV Clinical Research Centers. PATIENTS: Two-hundred and five patients with less than 4 weeks antiretroviral treatment, naive to 3TC and protease inhibitors and with CD4 cell counts > or = 200 x 10(6)/l and plasma HIV-1 RNA levels > or = 10,000 copies/ml. INTERVENTIONS: Stavudine 40 mg and ddI 200 mg twice daily plus IDV 800 mg every 8 h compared with ZDV 200 mg every 8 h or 300 mg twice daily, 3TC 150 mg twice daily plus IDV. MAIN OUTCOME MEASURES: The proportion of patients with plasma HIV-1 RNA levels < 500 copies/ml and < or = 50 copies/ml and changes in CD4 cell counts were compared. RESULTS: In an analysis of the primary endpoint, 61% of patients on d4T + ddI + IDV and 45% of patients on ZDV + 3TC + IDV had all HIV-1 RNA values obtained between weeks 40 and 48 < 500 copies/ml [95% confidence interval (CI) for the difference between proportions, 1.7-30.3%; P = 0.038]. In an intent-to-treat analysis, the percentage of all patients randomized with all HIV-1 RNA levels < 500 copies/ml between 40 and 48 weeks were 53% for the d4T + ddI + IDV arm and 41% for the ZDV + 3TC + IDV arm (95% CI, -1.4% to 25.7%; P = 0.068). At 48 weeks 41% and 35% were < or = 50 copies/ml for the stavudine- and ZDV-containing arms respectively (P > 0.2). The median time-weighted average increases in CD4 cells count over 48 weeks were 150 x 10(6)/l cells for the d4T arm and 106 x 10(6)/l cells for the ZDV arm (P= 0.001). The occurrence of serious adverse events was not significantly different between arms. CONCLUSION: The combination of stavudine, ddl and IDV resulted in potent antiretroviral effects over a 48-week period, comparable or superior to zidovudine, 3TC and IDV supporting the use of stavudine, ddI and a protease inhibitor as an initial antiretroviral treatment.

p27/kip1 expression in oligodendrogliomas and its possible prognostic role.

p27/kip1 regulates the G1-S transition of the cell cycle by inhibiting cyclinD-CDK4, cyclinE-CDK2 and cyclinA-CDK2 complexes. Regulation of p27 levels occurs mainly post-translationally by ubiquitin-mediated proteasomal proteolysis. Although genetic changes of p27/kip1 are extremely rare, in many human carcinomas p27 levels are reduced, correlate with histological malignancy, and are associated with poor prognosis. In astrocytic gliomas, p27 decreases with anaplasia and is almost absent in glioblastomas. p27/kip1 was immunohistochemically studied in 37 oligodendrogliomas, categorized according to WHO classification. In this series, the immunohistochemical reaction for p27 was confined to nuclei. p27 score showed a tendency to decrease with malignancy. When the p27 score was considered as high versus low expression (cut-off of p27 labeling index, LI, at 25%), it represented an independent prognostic factor in univariate (P = 0.02) and in multivariate analysis (P = 0.04). The risk ratio suggested that the p27 low expression group had a threefold increased possibility to show a reduced survival. Moreover, p27 levels did not correlate with MIB-1 LI, suggesting that p27 is not merely associated with the control of proliferation.

Prognostic significance of telomerase in brain tumors

Telomeres help to maintain chromosomal stability and integrity. They are progressively lost, conditioning aging and mortality. Telomerase enzyme activation stabilizes the telomere length, permitting the proliferation of cancer cells. Four papers are reviewed dealing with increased expression of telomerase associated with increased labelling index of MIB-1 (Ki-67) and malignancy in astrocytomas, ependymomas, and oligodendrogliomas. Telomerase expression can be utilized for differentiating malignant from benign tumors with histological resemblance. The significance of telomerase in tumor progression is discussed within the field of brain neoplasias.

Cyclin D1 expression in gliomas.

Cyclin D1 (cycD1) expression was defined immunohistochemically using monoclonal antibody DCS-6 and polyclonal antiserum H-295 in 50 glioma biopsies. The number of positive nuclei was higher for H-295 than for DCS-6, with a ratio of 3:1. The labelling index (LI) was compared to the grade of histological malignancy and to Ki-67 MIB-1 LI. The LI for cycD1 increased with histological malignancy, in parallel with the increase in MIB-1 LI. In most tumours, the maximum LI for cycD1 and MIB-1 were found in the same areas. The mean MIB-1 LI: mean cycD1 LI ratio does not vary in the three grades of astrocytic tumours. However, in this study the correlation between the two LIs was not statistically significant. Staining for cycD1 antigen does not necessarily imply that the gene is overexpressed since other molecular mechanisms can also be responsible for cell cycle deregulation. In invasive areas, the cycD1 LI is frequently higher than in solid tumour, either because more tumour cells are positive or because reactive astrocytes and activated microglia express cycD1. The relative contribution of neoplastic and reactive cells remains to be defined.

CDKN2/p16 inactivation and p16 immunohistochemistry in astrocytic gliomas.

CDKN2/p16 inactivation is the most frequent alteration in the molecular regulation of G1-S transition. CDKN2/p16 homozygous deletions was studied in paraffin-embedded sections of 45 astrocytic tumours by multiplex PCR. Immunohistochemistry for p16 and proliferation marker Ki-67 MIB-1 was performed in adjacent sections; their labelling index (LI) have been calculated. CDKN2/p16 gene was not deleted in astrocytomas, while homozygous deletion was found in 26.7% anaplastic astrocytomas, and in 55.0% of glioblastomas. Analysis of CDKN2/p16 homozygous deletion in discrete areas of the same tumour, showed that the deletion occurred independently of the phenotypic aspect of the areas. Nevertheless a genotypic and phenotypic heterogeneity is present in few cases. p16 immunohistochemistry mostly corresponds to the genotypic pattern. No correlation was found between CDKN2/p16 homozygous deletion and MIB-1 LI.

[Chronic gallbladder disease & its surgical therapy, personal experience]. / Incidenza della colecistopatia cronica e suo trattamento chirurgico: esperienza personale.

The chronic alitiasic cholecystopathy represents an unusual trmorphological disease not well defined in literature; it is characterized by a chronic alteration of gallbladder without gallstones. The authors present a retrospective study on a group of 21 patients having undergone cholecistectomy from January 1992 to January 1995. The choice for the surgical treatment was made on the basis of blind gallbladder after i.v. cholangiography, and cholecystography and/or tickning of the gallbladder wall performed through ultrasonography, in patients with hystory of biliary pains not responding to the pharmacological therapy. The removed gallbladder has been histologically examined, observing the absence of gallstone and the presence of chronic inflammation of the gallbladder wall. The c.a.c. is a pathology whose etiopatogenesis is yet discussed, some authors assert in fact that on its basis there are functional symptoms. Although in the past these kinds of disease were considered functional disorders this study revealed that in some cases there are organic problems and that the surgical treatment is necessary.

[Plasma levels of the serum antioxidants (uric acid, ceruloplasmin, transferrin) in term and preterm neonates in the first week of life]. / Livelli plasmatici degli antiossidanti sierici (acido urico, ceruloplasmina, transferrina) in neonati a termine e pretermine nella prima settimana di vita.

Development of serum antioxidants (ceruloplasmin, transferrin, uric acid and bilirubin) an the 1st, 4th, 7th days of life has been evaluated in 50 healthy NGA newborns (25 preterm with 35 +/- 0.6 week of gestational age and 2270 +/- 150 g of weight, 25 a term with 39 +/- 0.8 week of gestational age and 3480 +/- 220 g of weight) and in apparent absence of oxidant stress. The ceruloplasmin values increase from the 1st to 7th days of life and change significantly at the 4th day between a term and preterm newborns (p < 0.01); the transferrin values reduce significantly an the 1st (p < 0.01) and 7th days of life (p < 0.05); the uric acid values reduce in the two groups an the 1st day (p < 0.01). Our results show in the newborn a prevalent antioxidant activity of the studied substances. The plasma levels of uric acid may be compared, in the first week of life, to the hypoxanthine levels as acute ipoxia gauges.

Selective inhibition of cholesterol synthesis in liver versus extrahepatic tissues by HMG-CoA reductase inhibitors.

Hepatic specificity of inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase may be achieved by efficient first-pass liver extraction resulting in low circulating drug levels, as with lovastatin, or by lower cellular uptake in peripheral tissues, seen with pravastatin. BMY-21950 and its lactone form BMY-22089, new synthetic inhibitors of HMG-CoA reductase, were compared with the major reference agent lovastatin and with the synthetic inhibitor fluindostatin in several in vitro and in vivo models of potency and tissue selectivity. The kinetic mechanism and the potency of BMY-21950 as a competitive inhibitor of isolated HMG-CoA reductase were comparable to the reference agents. The inhibitory potency (cholesterol synthesis assayed by 3H2O or [14C]acetate incorporation) of BMY-21950 in rat hepatocytes (IC50 = 21 nM) and dog liver slices (IC50 = 23 nM) equalled or exceeded the potencies of the reference agents. Hepatic cholesterol synthesis in vivo in rats was effectively inhibited by BMY-21950 and its lactone form BMY-22089 (ED50 = 0.1 mg/kg p.o.), but oral doses (20 mg/kg) that suppressed liver synthesis by 83-95% inhibited sterol synthesis by only 17-24% in the ileum. In contrast, equivalent doses of lovastatin markedly inhibited cholesterol synthesis in both organs. In tissue slices from rat ileum, cell dispersions from testes, adrenal, and spleen, and in bovine ocular lens epithelial cells, BMY-21950 inhibited sterol synthesis weakly in vitro with IC50 values 76- and 188-times higher than in hepatocytes; similar effects were seen for BMY-22089. However, the IC50 ratios (tissue/hepatocyte) for lovastatin and fluindostatin were near unity in these models. Thus, BMY-21950 and BMY-22089 are the first potent synthetic HMG-CoA reductase inhibitors that possess a very high degree of liver selectivity based upon differential inhibition sensitivities in tissues. This cellular uptake-based property of hepatic specificity of BMY-21950 and BMY-22089, also manifest in pravastatin, is biochemically distinct from the pharmacodynamic-based disposition of lovastatin, which along with fluindostatin exhibited potent inhibition in all tissues that were exposed to it.

[Glycerol erythrocyte lysis test in the 1st month of life in neonates at term and premature infants]. / Il test di lisi eritrocitaria al glicerolo nel primo mese di vita in neonati a termine e pretermine.

Foetal erythrocytes have an enhanced resistance to osmotic haemolysis that they retain for the first 5 days after birth, particularly in premature newborns. The erythrocyte fragility test was used to study the increased resistance to osmotic haemolysis in 155 healthy newborns (50 NGA born to term, 55 NGA premature and 50 SGA born to term) as well as 31 newborns (20 born to term and 11 premature) with aspecific hyperbilirubinaemia. The drug used was glycerol which is specific in its reaction to alterations in erythrocyte membrane stability and made it possible to assess changes arising in the first month after birth, both naturally and in response to phototherapy. The results show a much higher resistance in neonatal than adult erythrocytes that gradually decrease during the first month after birth without, however, falling to adult levels. No changes in resistance attributable to the phototherapy adopted in newborns with hyperbilirubinaemia were encountered. The glycerol test proved extremely sensitive in the diagnosis of neonatal haemolytic anaemias easy to use so that congenital spherocytosis can be identified earlier than is otherwise possible.

Postoperative acute gastrointestinal tract hemorrhage and multiple-organ failure.

Acute upper gastrointestinal tract hemorrhage (AUGH) was evaluated postoperatively in 720 critically ill patients and correlated with multiple-organ failure (MOF). The AUGH incidence was 20.1%. Patients were divided into three groups. Group 1 (n = 77) patients were admitted without AUGH, but developed MOF and later AUGH, with renal failure as the most common previous failure. Group 2 (n = 36) patients were admitted with AUGH and other failures. Group 3 (n = 32) patients were admitted without AUGH, which appeared as the first or only failure. Means +/- SDs for MOF and mortality for groups 1, 2, and 3, respectively, were as follows: 3.2 +/- 0.8, 75.3%; 3.2 +/- 1.1, 63.9%; and 1.8 +/- 0.8, 28.1%. A control group (n = 90) with MOF but without AUGH presented 1.8 +/- 0.9 for MOF and 41.1% mortality. Mortality, sepsis, and mean MOF were higher in AUGH cases and lower in group 3 vs groups 1 and 2. Acute upper gastrointestinal tract hemorrhage is a component of MOF (Baue's syndrome) that is closely related to sepsis particularly after abdominal surgery.

Hemorragia digestiva aguda alta postoperatoria en pacientes criticos tratados en una unidad de terapia intensiva. / Acute upper gastrointestinal hemorrhage in postoperative period in critical patients treated in an intensive care unit

Se presenta una experiencia, en la observacion y tratamiento de enfermos criticos con hemorragia digestiva aguda alta postoperatoria. Se estudia la correlacion entre esta complicacion y el sindrome de insuficiencia multiple de organos y sistemas.Sobre 247 enfermos quirurgicos, se observaron 49 hemorragias (19,8%), con una mortalidad del 63,3%. Presentaron complicaciones septicas 40 pacientes (81,6%); fueron tratados quirurgicamente en todos los casos, sus focos septicos. Cursaron con el sindrome el 69,4% de los enfermos. Con el analisis retrospectivo de la experiencia se comprueba la importancia fundamental de a) la prevencion del sindrome, b) la deteccion precoz del mismo, c) el tratamiento electivo y temprano, de cada una de las insuficiencias organicas, evitando con ello la incorporacion al cuadro de otros parenquimas y d) ante la comprobacion de esta hemorragia es obligatorio buscar un foco septico, descartar la insuficiencia de otros organos y sistemas y no someter al paciente a cirugia directa de la hemorragia, sin antes descartar la existencia del sindrome

Hemorragia digestiva aguda alta postoperatoria en pacientes criticos tratados en una unidad de terapia intensiva. / Acute upper gastrointestinal hemorrhage in postoperative period in critical patients treated in an intensive care unit

Se presenta una experiencia, en la observacion y tratamiento de enfermos criticos con hemorragia digestiva aguda alta postoperatoria. Se estudia la correlacion entre esta complicacion y el sindrome de insuficiencia multiple de organos y sistemas.Sobre 247 enfermos quirurgicos, se observaron 49 hemorragias (19,8%), con una mortalidad del 63,3%. Presentaron complicaciones septicas 40 pacientes (81,6%); fueron tratados quirurgicamente en todos los casos, sus focos septicos. Cursaron con el sindrome el 69,4% de los enfermos. Con el analisis retrospectivo de la experiencia se comprueba la importancia fundamental de a) la prevencion del sindrome, b) la deteccion precoz del mismo, c) el tratamiento electivo y temprano, de cada una de las insuficiencias organicas, evitando con ello la incorporacion al cuadro de otros parenquimas y d) ante la comprobacion de esta hemorragia es obligatorio buscar un foco septico, descartar la insuficiencia de otros organos y sistemas y no someter al paciente a cirugia directa de la hemorragia, sin antes descartar la existencia del sindrome