Tree Growth and Climate Relationship : dynamics of Scots Pine (Pinus Sylvestris L.) Growing in the Near-Source Region of the Combined Heat and Power Plant During the Development of the Pro-Ecological Strategy in Poland

Since the 1990s, the emission of pollutants was reduced in a majority of Polish and developing country factories whereas the level of energy production was similar to that prior to the 1990s. The conifer investigated in this study has grown for many years under the stress of industrial pollution. Despite this, the trees are preserved, to a large extent, sensitive to the natural climatic factors. We present a complex analysis of the climatic (sunshine, temperature, precipitation, humidity, and wind circulation) and anthropogenic factors influencing the radial increment dynamics of Scots pine (Pinus sylvestris L.) growing in the vicinity of the combined heat and power station in Łaziska (Poland). We analyzed the spatiotemporal distribution of growth reductions, the depth of reduction with respect to the distance from the emitter, the relationship between tree growth and climate during the industry development period and during proecological strategy application . Samples of carbon isotopic composition in pine needles from 2012 to 2013 were additionally determined. Pines series of 3 positions indicate that they have a similar sensitivity to most climatic elements of the previous and given year, but there is also a different rhythm between the studied populations of incremental growth of pines. The causes of diversity are due to the different types of habitat (site types) and industrial pollution. The variation in carbon stable isotopic composition in pine needles was connected with an increase of CO2

Purification of phage display-modified bacteriophage T4 by affinity chromatography

<p>Abstract</p> <p>Background</p> <p>Affinity chromatography is one of the most efficient protein purification strategies. This technique comprises a one-step procedure with a purification level in the order of several thousand-fold, adaptable for various proteins, differentiated in their size, shape, charge, and other properties. The aim of this work was to verify the possibility of applying affinity chromatography in bacteriophage purification, with the perspective of therapeutic purposes. T4 is a large, icosahedral phage that may serve as an efficient display platform for foreign peptides or proteins. Here we propose a new method of T4 phage purification by affinity chromatography after its modification with affinity tags (GST and Histag) by <it>in vivo </it>phage display. As any permanent introduction of extraneous DNA into a phage genome is strongly unfavourable for medical purposes, integration of foreign motifs with the phage genome was not applied. The phage was propagated in bacteria expressing fusions of the phage protein Hoc with affinity tags from bacterial plasmids, independently from the phage expression system.</p> <p>Results</p> <p>Elution profiles of phages modified with the specific affinity motifs (compared to non-specific phages) document their binding to the affinity resins and effective elution with standard competitive agents. Non-specific binding was also observed, but was 10²-10⁵times weaker than the specific one. GST-modified bacteriophages were also effectively released from glutathione Sepharose by proteolytic cleavage. The possibility of proteolytic release was designed at the stage of expression vector construction. Decrease in LPS content in phage preparations was dependent on the washing intensity; intensive washing resulted in preparations of 11-40 EU/ml.</p> <p>Conclusions</p> <p>Affinity tags can be successfully incorporated into the T4 phage capsid by the <it>in vivo </it>phage display technique and they strongly elevate bacteriophage affinity to a specific resin. Affinity chromatography can be considered as a new phage purification method, appropriate for further investigations and development.</p

Analysis of postural disorders using a stabilometric platform in patients with rheumatic diseases

Wstęp W przebiegu reumatoidalnego zapalenia stawów (RZS) ważnym problemem wpływającym na stabilność oraz symetrię obciążenia kończyn dolnych jest globalne upośledzenie motoryki stopy spowodowane jej płasko-koślawą deformacją, a także występowanie licznych zmian w obrębie stawów stóp. W zesztywniającym zapaleniu stawów kręgosłupa (ZZSK) dochodzi ostatecznie do trwałego ograniczenia ruchomości kręgosłupa co prowadzi do zaburzeń prawidłowego obciążania kończyn dolnych i równowagi ciała. W twardzinie układowej (TU) ból i przykurcze stawowe negatywnie oddziałują na prawidłową postawę i stabilność ciała oraz prawidłowe obciążenie kończyn dolnych. Metodyka Badanie balansu ciała oraz stabilności i symetrii obciążenia kończyn dolnych wykonano na komputerowej dwupłytowej platformie stabilometrycznej. Badanie składało się z dwóch prób – pierwszej z otwartymi i drugiej z zamkniętymi oczami. Wyniki Ogółem przebadano 50 chorych z ZZSK, 68 chorych z RZS oraz 53 chorych z TU. Nie wykazano istotnych statystycznie różnic między badanymi grupami z uwzględnieniem badania z oczami otwartymi jak i zamkniętymi. W każdej grupie badanej wykazano istotnie statystycznie wyższe wartości całkowitej drogi, którą przebył środek nacisku stóp chorego w ciągu badania, ocenianego przy zamkniętych oczach względem ocenianego przy otwartych oczach. Dyskusja Analizowane w pracy grupy stanowią trzy odrębne funkcjonalnie choroby tkanki łącznej, charakteryzujące się zajęciem innych składowych elementu ruchu. Twardzina układowa jawi się jako choroba o zupełnie odmiennym wzorcu ruchu charakteryzującego się zmniejszeniem wychylenia środka nacisku stóp zarówno w badaniu z zamkniętymi jak i otwartymi oczami. Wnioski W chorobach reumatycznych powszechnie występują zaburzenia postawy i prawidłowego utrzymania równowagi, a wzorzec zaburzeń ruchowych jest zależny od danej choroby.Background: In the course of rheumatoid arthritis (RA), an important problem that affects the stability and symmetry of lower limb loading is the global motor impairment of the foot due to its planovalgus deformity, as well as the occurrence of numerous lesions in the foot joints. In ankylosing spondylitis (AS), there is eventually a permanent limitation of spinal mobility, which results in impaired lower limb loading and body balance. In systemic sclerosis (SSc), pain and joint contractures negatively affect the proper posture and stability of the body and the normal lower limb loading. Material and methods:Tests of body balance, stability and symmetry of lower limb loading were performed on a computerised two-plate stabilometric platform. The testing consisted of two trials — the first performed with eyes open (open-eye test) and the second performed with eyes closed (closed-eye test). Results: There were a total of 50 AS patients, 68 RA patients and 53 SSc patients examined. There were no statistically significant differences between the study groups including the open-eye test and the closed-eye test. Each study group revealed statistically significant higher values for the total distance travelled by the patient's centre of pressure of the feet over the course of testing as assessed using the closed-eye test compared to the open-eye test. Discussion: The groups analysed in this paper represent three functionally distinct connective tissue diseases that are marked by involvement of different constituents of the motor system component. Systemic sclerosis (SSc) presents itself as a disease with a completely different movement pattern that is marked by a reduction in the amplitudes of the centre of pressure (CoP) in both the closed-eye test and the open-eye test. Conclusions: Postural and balance disorders are common in rheumatic diseases, and the pattern of movement disorders is disease-specific.

The effect of bacteriophages T4 and HAP1 on in vitro melanoma migration

<p>Abstract</p> <p>Background</p> <p>The antibacterial activity of bacteriophages has been described rather well. However, knowledge about the direct interactions of bacteriophages with mammalian organisms and their other, i.e. non-antibacterial, activities in mammalian systems is quite scarce. It must be emphasised that bacteriophages are natural parasites of bacteria, which in turn are parasites or symbionts of mammals (including humans). Bacteriophages are constantly present in mammalian bodies and the environment in great amounts. On the other hand, the perspective of the possible use of bacteriophage preparations for antibacterial therapies in cancer patients generates a substantial need to investigate the effects of phages on cancer processes.</p> <p>Results</p> <p>In these studies the migration of human and mouse melanoma on fibronectin was inhibited by purified T4 and HAP1 bacteriophage preparations. The migration of human melanoma was also inhibited by the HAP1 phage preparation on matrigel. No response of either melanoma cell line to lipopolysaccharide was observed. Therefore the effect of the phage preparations cannot be attributed to lipopolysaccharide. No differences in the effects of T4 and HAP1 on melanoma migration were observed.</p> <p>Conclusion</p> <p>We believe that these observations are of importance for any further attempts to use bacteriophage preparations in antibacterial treatment. The risk of antibiotic-resistant hospital infections strongly affects cancer patients and these results suggest the possibility of beneficial phage treatment. We also believe that they will contribute to the general understanding of bacteriophage biology, as bacteriophages, extremely ubiquitous entities, are in permanent contact with human organisms.</p

The Effects of Duodenojejunal Omega Switch in Combination with High-Fat Diet and Control Diet on Incretins, Body Weight, and Glucose Tolerance in Sprague-Dawley Rats

Background: Despite excellent results of bariatric surgery in the treatment of type 2 diabetes and weight loss in human subjects, some patients do not obtain desired results. One of the reasons for this is that not all patients follow caloric intake recommendations. Aim: The aim of this study was to investigate the effect of duodenojejunal omega switch (DJOS) surgery on body weight, glucose tolerance, and incretins in rats. Methods: DJOS and SHAM surgery were performed on rats maintained for 8 weeks on high-fat diet (HF) and control diet (CD), respectively. After surgery, four groups were kept on the same diet as before the surgery, and four groups had a changed diet (CD vs. HF and HF vs. CD) for the next 8 weeks. Glucose tolerance, insulin tolerance, glucose-stimulated insulin, glucagon-like peptide-1 (GLP-1) and gastric inhibitory polypeptide/glucose-dependent insulinotropic polypeptide (GIP) secretion, food intake, and body weight were measured. Results: A change of diet after surgery resulted in reduced glucose tolerance. Plasma insulin levels were lowered between DJOS and SHAM surgeries for the HF/HF and CD/HF groups. DJOS surgery did not reduce body weight in the studied groups, irrespective of diet. In the HF/HF group, ΔGLP-1 was lower for DJOS surgery in comparison with other groups. Differences of weight changes were observed for groups HF/HF and HF/CD. After DJOS surgery, ΔGIP was lower in the CD/HF group compared with HF/HF. Conclusions: Our results show that applications of different types of diets, before and after surgery, is a sensitive method for studies of mechanism of glucose intolerance after DJOS surgery

Wpływ zmiany wytycznych ASCO-CAP na ocenę statusu genu HER2 metodą FISH w kwalifikacji do terapii anty-HER2 w raku piersi

Introduction. Breast cancer is the most common cancer among Polish women. Overexpression of the HER2 protein or HER2 gene amplification is associated with a poor prognosis, simultaneously being an indication for the HER2-targeted therapy. In equivocal cases, the FISH assay is used for the final identification of the HER2 gene status. This evaluation should be performed according to the ASCO-CAP guidelines which have been changed in 2013. The aim of this study was to assess whether and how the changes of recommendations affected the distribution of the FISH results. Materials and methods. The results of routine diagnostic FISH analyses were compared for two independent groups of patients assessed with different evaluation criteria (ASCO-CAP 2007 for n = 680 and ASCO-CAP 2013 n = 851), and also in a group of 763 patients, where both criteria were used simultaneously. Results. A comparison of the results obtained in two independent groups showed that the change of evaluation criteria did not alter the percentage of HER2-positive tests (with HER2 amplification). However, the frequency of HER2-negative analyses (without HER2 amplification) diminished significantly from 76.2% to 61.8%, whereas the equivocal group (with an indefinite status of HER2 amplification) increased from 0.4% to 13.6%. In the group where both criteria from 2007 and 2013 were used, we also discovered statistically significant differences. The frequency of HER2-positive results were elevated from 10.6% to 16.8%. The equivocal results were also found more often, rising from 4.2% to 15.6%, while the number of negative results lowered from 85.2% to 67.6%. Conclusions. The use of ASCO/CAP recommendations for the assessment of the HER2 gene status reduces the group of negative results, and concurrently enlarges the number of positive and equivocal ones. This indicates that the new criteria extends the access to HER2-targeted therapy. Nevertheless, they also raise the frequency of analyses with an indefinite status of the HER2 gene. Our outcome suggests that there is a need for an enhanced FISH-based evaluation of this gene in the last group of patients in order to provide them with an unambiguous stratification to risk groups.  Wstęp. Rak piersi jest najczęstszym nowotworem u kobiet w Polsce. Nadmierna ekspresja białka HER2 lub amplifikacja genu HER2 jest związana ze złym rokowaniem i stanowi wskazanie do zastosowania terapii anty-HER2. W przypadkach wątpliwych rozstrzygającym badaniem jest ocena FISH, wskazująca status amplifikacji genu HER2 według obowiązujących wytycznych ASCO-CAP, które w roku 2013 uległy zmianom. Celem pracy było sprawdzenie, czy i w jaki sposób zmiana zaleceń wpłynęła na rozkład wyników badań FISH w tej grupie pacjentów. Materiały i metody. Analizie porównawczej poddano wyniki rutynowej diagnostyki metodą FISH w dwóch niezależnych grupach pacjentów z zastosowaniem dwóch różnych kryteriów oceny (ASCO-CAP 2007 dla n = 680 i ASCO-CAP 2013 dla n = 851) oraz w grupie 763 pacjentów, gdzie zastosowano równolegle obydwa kryteria oceny. Wyniki. Porównanie wyników uzyskanych w dwóch niezależnych grupach wykazało brak istotnej zmiany odsetka wyników HER2-dodatnich (z amplifikacją) po zmianie kryteriów oceny. Istotne statystycznie okazało się zmniejszenie grupy wyników HER2-negatywnych (bez amplifikacji) z 76,2% na 61,8% przy rozszerzeniu grupy niejednoznacznej (o nieokreślonym statusie amplifikacji) z 0,4% do 13,6%. Grupa badana równolegle wg kryteriów 2007 i 2013 wykazała różnice istotne statystycznie. Zanotowano wzrost przypadków HER2-pozytywnych z 10,6% do 16,8%, znaczny wzrost wyników niejednoznacznych, z 4,2% na 15,6%, przy równoczesnym spadku wyników negatywnych z 85,2% na 67,6%. Wnioski. Zastosowanie nowych wytycznych ASCO-CAP 2013 w ocenie statusu genu HER2 wpływa na zawężenie grupy wyników negatywnych, natomiast rozszerza grupę wyników pozytywnych i niejednoznacznych. Wynik taki wskazuje na rozszerzenie dostępu do kwalifikacji w kierunku terapii anty-HER2. Natomiast istotny wzrost odsetka pacjentów z wynikiem o nieokreślonym statusie amplifikacji genu HER2 wskazuje na konieczność pogłębionej w tej grupie oceny FISH w celu uzyskania możliwości jednoznacznej stratyfikacji do grup ryzyka.

Dual Mechanism of Interleukin-3 Receptor Blockade by an Anti-Cancer Antibody

SummaryInterleukin-3 (IL-3) is an activated T cell product that bridges innate and adaptive immunity and contributes to several immunopathologies. Here, we report the crystal structure of the IL-3 receptor α chain (IL3Rα) in complex with the anti-leukemia antibody CSL362 that reveals the N-terminal domain (NTD), a domain also present in the granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-5, and IL-13 receptors, adopting unique “open” and classical “closed” conformations. Although extensive mutational analyses of the NTD epitope of CSL362 show minor overlap with the IL-3 binding site, CSL362 only inhibits IL-3 binding to the closed conformation, indicating alternative mechanisms for blocking IL-3 signaling. Significantly, whereas “open-like” IL3Rα mutants can simultaneously bind IL-3 and CSL362, CSL362 still prevents the assembly of a higher-order IL-3 receptor-signaling complex. The discovery of open forms of cytokine receptors provides the framework for development of potent antibodies that can achieve a “double hit” cytokine receptor blockade

Immunogenic epitope scanning in bacteriolytic enzymes Pal and Cpl-1 and engineering Pal to escape antibody responses

Bacteriolytic enzymes are promising antibacterial agents, but they can cause a typical immune response in vivo. In this study, we used a targeted modification method for two antibacterial endolysins, Pal and Cpl-1. We identified the key immunogenic amino acids, and designed and tested new, bacteriolytic variants with altered immunogenicity. One new variant of Pal (257-259 MKS → TFG) demonstrated decreased immunogenicity while a similar mutant (257-259 MKS → TFK) demonstrated increased immunogenicity. A third variant (280-282 DKP → GGA) demonstrated significantly increased antibacterial activity and it was not cross-neutralized by antibodies induced by the wild-type enzyme. We propose this variant as a new engineered endolysin with increased antibacterial activity that is capable of escaping cross-neutralization by antibodies induced by wild-type Pal. We show that efficient antibacterial enzymes that avoid cross-neutralization by IgG can be developed by epitope scanning, in silico design, and substitutions of identified key amino acids with a high rate of success. Importantly, this universal approach can be applied to many proteins beyond endolysins and has the potential for design of numerous biological drugs

Circulation of Fluorescently Labelled Phage in a Murine Model

Interactions between bacteriophages and mammals strongly affect possible applications of bacteriophages. This has created a need for tools that facilitate studies of phage circulation and deposition in tissues. Here, we propose red fluorescent protein (RFP)-labelled E. coli lytic phages as a new tool for the investigation of phage interactions with cells and tissues. The interaction of RFP-labelled phages with living eukaryotic cells (macrophages) was visualized after 20 min of co-incubation. RFP-labeled phages were applied in a murine model of phage circulation in vivo. Phages administered by three different routes (intravenously, orally, rectally) were detected through the course of time. The intravenous route of administration was the most efficient for phage delivery to multiple body compartments: 20 min after administration, virions were detected in lymph nodes, lungs, and liver; 30 min after administration, they were detectable in muscles; and 1 h after administration, phages were detected in spleen and lymph nodes. Oral and rectal administration of RFP-labelled phages allowed for their detection in the gastrointestinal (GI) tract only