Crude liver membrane fractions as substrate preserve liver-specific functions in long-term,serum-free rat hepatocyte cultures

Summary Over time, rat hepatocytes cultured on collagen lose the capacity to express liver-specific functions. The influence on this degradation process of an alternative substratum—crude membrane fractions prepared from the liver of the same rat strain—was investigated. Freshly isolated rat hepatocytes were cultured in serum-free Williams E medium supplemented with aprotinin, selenium, dexamethasone, and insulin in flasks coated with a mixture of rat liver crude membrane fractions:collagen type I (100:1). The cells adhered firmly, exhibiting minimal spreading and remaining grouped in columns or in cell islands, and retained their liver-specific functions for more than 1 wk. Hepatocytes secreted substantially higher amounts of albumin than cells cultured on collagen-coated dishes, and on Days 1 and 9 in culture the total P-450 content was 72 and 40%, respectively, of that of freshly isolated cells. On Day 6, the 7-ethoxyresorufin-O-de-ethylase and the aldrin epoxidase activities were still more than 50% that of freshly isolated hepatocytes. Exposure to phenobarbital on Days 3 to 6 increased the total cytochrome P-450 content twofold; exposure to 3-methylcholanthrene increased the activity of the corresponding cytochrome P-450 isoforms to 20 times that observed in untreated cultures and 6 times that observed in freshly isolated cells. Thus, given the ease with which they are prepared, the use of crude membrane fractions combined with culture medium supplemented with aprotinin and selenium can facilitate the preparation of reproducible cultures suitable for long-term in vitro pharmacotoxicologic studies using rat hepatocytes.     

A study on duplications of the dystrophin gene: Evidence of a geographical difference in the distribution of breakpoints by intron

Starting from a group of 265 Italian patients affected with Duchenne or Becker muscular dystrophy a screening for duplications in the dystrophin gene was performed on 112 cases in which no deletions had previously been detected. The 21 intragenic duplications detected account for 7.9% of the total. Among these, one duplication including exons from 3 to 43 is the largest reported so far. Data from this study were combined with those from the literature and breakpoint distribution by intron was analysed. In general breakpoints occur mostly in the proximal third of the gene, in particular in intron 7. However, both the frequency of duplications and the distribution of breakpoints by intron are different in the Japanese sample compared with the other groups of patients. The role of geographical differentiation of intron sequences by genetic drift and of transposon-like sequences in explaining these differences is discussed.     

The importance of hornbeam (Carpinus sp.) pollen hypersensitivity in spring allergies

Hornbeam (Carpinus sp.) flowering occurs generallysimultaneously with birch anthesis. While birch pollenis known as the major tree-allergen in central andnorthern Europe, the quantitative and qualitative roleof hornbeam pollen in allergy has not been evaluated.In some regions of Switzerland, the annual ratiohornbeam SPI / birch SPI reaches 20–30% on average,and 50–180% some years. The relative importance ofhornbeam pollen hypersensitivity has been evaluated inmore than 200 patients suffering from spring allergiesto pollen during the last five years (1993–1997) inthe region of Geneva. Among symptomatic patientsduring the March-April period, 73.5% werehypersensitive to Betulaceae pollen. Overall, 71.7%of the patients allergic to Betulaceae pollen werefound to be sensitive to hornbeam, and 12.5% of thesame group of patients were sensitive to hornbeampollen only. Among this last group, half of thepatients, besides allergic rhino-conjunctivitis, werealso suffering from asthma. Hornbeam pollen isabundant and appears to have a significativeimportance in spring allergies. In some cases,specific immunotherapy to hornbeam pollen shouldperhaps be considered.     

Validated spectrofluorimetric and resonance Rayleigh scattering methods for determining naftidrofuryl in varied pharmaceutical samples based on its interaction with erythrosin B

Naftidrofuryl is a vasodilator medication used for treating cerebral and peripheral vascular diseases. In this study, two spectroscopical techniques, spectrofluorimetric and resonance Rayleigh scattering (RRS), were utilized to quantify naftidrofuryl in its pharmaceutical samples. The developed methodologies in this study rely on a facile process of forming an association complex between erythrosine B reagent and naftidrofuryl under acidic conditions. The fluorimetric assay is based on the ability of naftidrofuryl to quench and decrease the native fluorescence intensity of the reagent when measured at ${\lambda }_{\mathit{\text{emis}}.}$ = 550 nm ( ${\lambda }_{\mathit{\text{excit}}.}$ = 526 nm). Under similar reaction conditions, the RRS method relies on the observed amplification in the RRS spectrum of the reagent at a wavelength of 577 nm following its interaction with naftidrofuryl. The methods exhibited linearity within the ranges 0.2–1.6 μg/ml (r² = 0.999) and 0.1–1.4 μg/ml (r² = 0.9994), with limit of quantitation values of 0.146 and 0.099 μg/ml, and limit of detection values of 0.048 and 0.032 μg/ml, for the fluorometric and the RRS methods, respectively. Moreover, the quenching between the dye and naftidrofuryl was studied using Stern–Volmer analysis, and the methodologies were experimentally optimized and validated. Additionally, acceptable recoveries were achieved when the procedures were applied to determine naftidrofuryl in pharmaceutical samples.     

On the Approximation of Optimal Structures for RNA-RNA Interaction

The interaction of two RNA molecules is a common mechanism for many biological processes. Small interfering RNAs represent a simple example of such an interaction. But other more elaborate instances of RNA-RNA interaction exist. Therefore, algorithms that predict the structure of the RNA complex thus formed are of great interest. Most of the proposed algorithms are based on dynamic programming. RNA-RNA interaction is generally NP-complete; therefore, these algorithms (and other polynomial time algorithms for that matter) are not expected to produce optimal structures. Our goal is to characterize this suboptimality. We demonstrate the existence of constant factor approximation algorithms that are based on dynamic programming. In particular, we describe 1/2 and 2/3 factor approximation algorithms. We define an entangler and prove that 2/3 is a theoretical upper bound on the approximation factor of algorithms that produce entangler-free solutions, e.g., the mentioned dynamic programming algorithms.     

Assessing Treatment Benefit in Immuno-oncology

Immuno-oncology is a buoyant field of research, with recently developed drugs showing unprecedented response rates and/or a hope for a meaningful prolongat     

Effect of neurogenic acupoint cupping on high sensitive C-reactive protein and pain perception in female chronic pelvic pain: A randomized controlled trial

Objectives:To determine the effect of neurogenic acupoint dry cupping therapy on high sensitive C-reactive protein (hs-CRP) level, pain perception & intensity, and life impact of pelvic pain in women with chronic pelvic pain (CPP), with regard to the biological and neurophysiological impacts of dry cupping on acupoint.Methods:Thirty women with CPP were randomly divided into two equal groups; the study group received dry cupping on neurogenic acupoints plus lifestyle modifications for 8 weeks (n=15), while the control group received only lifestyle modifications for 8 weeks (n=15). Women were assessed pre- and post-rehabilitation program with the hs-CRP blood test, the short-form McGill Pain Questionnaire (SF-MPQ), and the pelvic pain impact questionnaire (PPIQ).Results:Comparing both groups post-treatment revealed that there were significant reductions in levels of hs-CRP, and scores of SF-MPQ & PPIQ (p<0.05) in the study group compared with the control group. Also, there were significant positive correlations between hs-CRP and both SF-MPQ “Visual Analogue Scale (VAS), Present Pain Intensity (PPI) index & Pain Rating Index (PRI)” and PPIQ (p<0.05).Conclusion:Neurogenic acupoint cupping therapy had significantly improving effects on the degree of inflammation, pain perception & intensity, and life impact of pelvic pain in women with CPP.     

Characterization of glycerol kinase and NAD-independent glycerol-3-phosphate dehydrogenase from Pediococcus pentosaceus N5p

The polymerase chain reaction (PCR) has the potential to detect low levels of the human pathogen Escherichia coli O157 : H7 in bovine faeces. To improve the utility of PCR for this application, several methods for preparing template DNA from bovine faeces, both directly and after non-selective enrichment, were tested. These were boiling, enzyme treatment, enzyme treatment plus phenol-chloroform extraction, and enzyme treatment plus phenol-chloroform extraction plus Geneclean^® purification. Of these, the boiling method was the most consistent and had a sensitivity of approximately 3 cfu g⁻¹ faeces, with an assay time of less than 32 h. The boiling method was also combined with immunomagnetic separation (IMS) to detect E. coli O157 : H7 in less than 8 h, but with a sensitivity of approximately 10³ cfu g⁻¹ faeces. These methods can be used to prepare template for PCR screening of bovine faeces using any appropriate PCR primers.