Stimulated Raman scattering microscopy in chemistry and life science - Development, innovation, perspectives.

In this review, we present a summary of the basics of the Stimulated Raman Scattering (SRS) phenomenon, methods of detecting the signal, and collection of the SRS images. We demonstrate the advantages of SRS imaging, and recent developments, but also the limitations, especially in image capture speeds and spatial resolution. We also compare the use of SRS microscopy in biological system studies with other techniques such as fluorescence microscopy, second-harmonic generation (SHG)-based microscopy, coherent anti-Stokes Raman scattering (CARS), and spontaneous Raman, and we show the compatibility of SRS-based systems with other discussed methods. The review is also focused on indicating innovations in SRS microscopy, on the background of which we present the layout and performance of our homemade setup built from commercially available elements enabling for imaging of the molecular structure of single cells over the spectral range of 800-3600 cm-1. Methods of image analysis are discussed, including machine learning methods for obtaining images of the distribution of selected molecules and for the detection of pathological lesions in tissues or malignant cells in the context of clinical diagnosis of a wide range of diseases with the use of SRS microscopy. Finally, perspectives for the development of SRS microscopy are proposed.

Effects of body temperature on post-anoxic oxidative stress from the perspective of postnatal physiological adaptive processes in rats.

It is well known that decrease in body temperature provides protection to newborns subjected to anoxia/ischemia. We hypothesized that the normal body temperature of 33°C in neonatal rats (4°C below normal body temperature in adults) is in fact a preadaptation to protect CNS from anoxia and further reductions as well as elevations in temperature may be counterproductive. Our experiments aimed to examine the effect of changes in body temperature on oxidative stress development in newborn rats exposed to anoxia. Two-day-old Wistar rats were divided into 4 temperature groups: i. hypothermic at body temperature of 31°C, ii. maintaining physiological neonatal body temperature of 33°C, iii. forced to maintain hyperthermic temperature of 37°C, and i.v. forced to maintain hyperthermic temperature of 39°C. The temperature was controlled starting 15 minutes before and afterword during 10 minutes of anoxia as well as for 2 hours post-anoxia. Cerebral concentrations of lipid peroxidation products malondialdehyde (MDA) and conjugated dienes (CD) and the activities of antioxidant enzymes had been determined post mortem: immediately after anoxia was finished and 3, 7, and 14 days later. There were no post-anoxic changes in the concentration of MDA, CD and in antioxidant enzymes activity in newborn rats kept at their physiological body temperature of 33°C. In contrast, perinatal anoxia at body temperature elevated to 37°C or 39°C as well as under hypothermic conditions (31°C) intensified post-anoxic oxidative stress and depleted the antioxidant pool. Overall, these findings suggest that elevated body temperature (hyperthermia or fever), as well as exceeding cooling beyond the physiological level of body temperature of newborn rats, may extend perinatal anoxia-induced brain lesions. Our findings provide new insights into the role of body temperature in anoxic insult in vivo.

A five-year perspective on the situation of haemorrhagic fever with renal syndrome and status of the hantavirus reservoirs in Europe, 2005-2010.

Hantavirus infections are reported from many countries in Europe and with highly variable annual case numbers. In 2010, more than 2,000 human cases were reported in Germany, and numbers above the baseline have also been registered in other European countries. Depending on the virus type human infections are characterised by mild to severe forms of haemorrhagic fever with renal syndrome. The member laboratories of the European Network for diagnostics of Imported Viral Diseases present here an overview of the progression of human cases in the period from 2005 to 2010. Further we provide an update on the available diagnostic methods and endemic regions in their countries, with an emphasis on occurring virus types and reservoirs.

Prostaglandin-endoperoxide synthase genes COX1 and COX2 - novel modifiers of disease severity in cystic fibrosis patients.

Cystic fibrosis (CF) is one of the most common autosomal recessive diseases among Caucasians caused by a mutation in the CFTR gene. However, the clinical outcome of CF pulmonary disease varies remarkably even in patients with the same CFTR genotype. This has led to a search for genetic modifiers located outside the CFTR gene. The aim of this study was to evaluate the effect of functional variants in prostaglandin-endoperoxide synthase genes (COX1 and COX2) on the severity of lung disease in CF patients. To the best of our knowledge, it is the first time when analysis of COX1 and COX2 as potential CF modifiers is provided. The study included 94 CF patients homozygous for F508del mutation of CFTR. To compare their clinical condition, several parameters were recorded, e.g. a unique clinical score: disease severity status (DSS). To analyse the effect of non-CFTR genetic polymorphisms on the clinical course of CF patients, the whole coding region of COX1 and selected COX2 polymorphisms were analysed. Statistical analysis of genotype-phenotype associations revealed a relationship between the heterozygosity status of identified polymorphisms and better lung function. These results mainly concern COX2 polymorphisms: -765G>C and 8473T>C. The COX1 and COX2 polymorphisms reducing COX protein levels had a positive effect on all analysed clinical parameters. This suggests an important role of these genes as protective modifiers of pulmonary disease in CF patients, due to inhibition of arachidonic acid conversion into prostaglandins, which probably reduces the inflammatory process.

Stress-induced behaviour in adult and old rats: effects of neonatal asphyxia, body temperature and chelation of iron.

Perinatal asphyxia in mammals leads to iron accumulation in the brain, which results in delayed neurobehavioural disturbances, including impaired learning and abnormal alertness over their entire life span. The aim of this investigation was to verify our hypothesis that newborn rats, showing reduced normal body temperature, are protected against neurotoxicity of the asphyxia up to senescence. Alertness was studied in adult and old male Wistar rats after exposure to critical neonatal anoxia: (i) at physiological neonatal body temperature of 33 degrees C, (ii) at body temperature elevated to 37 degrees C, or (iii) at body temperature elevated to 39 degrees C (the thermal conditions remained unchanged both during anoxia and for 2 h postanoxia). To elucidate the effect of iron-dependent postanoxic oxidative damage to the brain, half of the group (iii) was injected with deferoxamine, a chelator of iron. Postanoxic behavioural disturbances were recorded in open-field, elevated plus-maze, and sudden silence tests when the rats reached the age of 12 and 24 months. Open-field stress-induced motor activity was reduced in rats subjected to neonatal anoxia under hyperthermic conditions. In contrast, these rats were hyperactive in the plus-maze test. Both the plus-maze and sudden silence tests show reduced alertness of these rats to external stimuli signalling potential dangers. The behavioural disturbances were prevented by body temperature of 33 degrees C and by administration of deferoxamine.

Seasonal changes in cryoprotectants concentrations in Helix pomatia snails.

Terrestrial snails are often exposed to freezing. Therefore, we investigated seasonal shifts in hemolymph concentrations of cryoprotectants such as glycerol and glucose. We also investigated whether summer acclimation to cold and short-day photoperiod induced synthesis of cryoprotectants in Helix pomatia snails. Concentrations of the both cryoprotectants were elevated in winter and reduced in summer. These changes, however, were not correlated with shifts in liver glycogen content. Summer acclimation to cold (5 degrees C) and short-day photoperiod evoked a selective increase in glycerol concentration. In conclusion, glycerol may play a role in adaptation of the snails to winter cold and glucose is rather unlikely to provide the cryoprotection.

Effect of temperature on postanoxic, potentially neurotoxic changes of plasma pH and free iron level in newborn rats.

In asphyxiated newborns, iron, released from heme and ferritin and deposited in the brain, contributes to neurodegeneration. Because hypothermia provides neuroprotection, newborn mammals, showing reduced body temperature, might avoid iron-mediated neurotoxicity. However, hypothermia leads to acidosis, which induces hyperferremia. Therefore, we decided to study the effects of body temperature on plasma pH and iron levels in newborn rats exposed to a critical anoxia. Rectal temperature was kept at 33 degrees C (typical of neonates), reduced by 2 degrees C, or elevated to a level typical of healthy (37 degrees C) or febrile (39 degrees C) adults. Arterial blood samples were collected at 0, 10, 20, 30, and 120 min postanoxia. Control samples were obtained from normoxic, temperature-matched neonates. Anoxia tolerance time decreased progressively at rectal temperatures exceeding 33 degrees C. Neither pH nor plasma iron were significantly affected by anoxia at 33 degrees C. Although hypothermia (31 degrees C) resulted in acidosis in normoxic rats, both pH and iron levels were hardly influenced by anoxia. However, acidosis and hyperferremia, proportional to body temperature, developed at 37 and 39 degrees C. In conclusion, reduced body temperature is likely to protect asphyxiated newborns against iron-mediated brain injury.

[A study evaluating the correlation between the phenotype and genotype among 65 cystic fibrosis patients]. / Próba oceny zaleznosci pomiedzy fenotypem a genotypem wsród 65 chorych na mukowiscydoze.

Among 65 CF diagnosed patients with both CFTR gene mutations known genotype-phenotype studies were performed. Correlation between pancreatic insufficiency and so called "severe mutations" was found. Respiratory tract symptoms do not seem to depend on one specific mutation as well as meconium ileus is not only limited to the group of patients with delta F508/delta F508 genotype. Some other genotype - clinical features correlation in CF patients are discussed.

[Ocular symptoms in internal carotid artery occlusion]. / Objawy oczne w niedroznosci tetnicy szyjnej wewnetrznej.

The internal carotid artery occlusion causing an insufficiency of the cerebral circulation manifests itself--besides the neurological symptoms--by disorders of the visual organ. One can distinguish among them early, functional symptoms in the form of transient unilateral hemianopia on the side opposite to the occluded carotid artery and the attacks of transient amblyopia. Signs of fixed ischaemia of the eye ball appear afterwards in the form of oculomotor and pupillary disturbances, neovascularization of the iris, disturbances of the IOP, central retinal artery occlusion, ischaemic optic atrophy. The authors emphasize the value of Doppler's diagnostic ultrasonography, a fast and non-invasive method of direct evaluation of the blood flow in the internal carotid artery system.

Correlations between DNCB skin hypersensitivity and leukocyte migration inhibition by DNCB-coated human erythrocytes.

Correlations between skin hypersensitivity to DNCB and LMI in presence of lyophilized, DNCB-coated human erythrocytes were evaluated. False positive results were found in 2 out of 25 (8%) non-sensitized donors and false negative--in 5 out of 21 (24%) sensitized patients. In follow-up evaluation after topical DNCB sensitization significant MI was usually seen about 3 weeks after sensitization and only in a part of patients after 6-9 weeks. Time limitations in the application of LMI test for evaluation of effects of immunosuppressive treatment are discussed.

[Progress in the early detection of inborn errors of metabolism]. / Postepy we wczesnym rozpoznawaniu genetycznie uwarunkowanych zaburzen metabolicznych.

The aim of present study was to evaluate the effectiveness of screening program for early detection of some metabolic errors in newborn population. The examinations included: early diagnostic of some amino acids and carbohydrates disturbances, cystic fibrosis and congenital hypothyreosis. Guthrie test and multidirectional urine screening test were used for the diagnostics of inborn errors in amino acids metabolism. Guthrie test for phenylalanine proved its high effectiveness and taking into account the relatively high frequency of phenylketonuria in our population this screening has been introduced as obligatory. The evaluation of pilot screening for tyrosinemia, homocystinuria and histidinemia in spite of no objections as to the tests themselves proved low frequency of these disorders in our country, sofar these tests have been abandoned. Multidirectional urine screening carried out in 6-8 weeks old infants allows for follow up control for some aminoacidopathies, and also for the detection of some transport metabolism and other metabolic errors. There is no doubt that screening tests for galactosemia should be carried out because of severe course of the disease and good results of its treatment. Problem to be discussed is the choice of screening procedure and age at which it should be performed. Cystic fibrosis being one of the most common disease in the group of metabolic disorders needs to be screened, because the detection allows for early introduction of complex palliative treatment. The comparative evaluation of three meconium tests for cystic fibrosis revealed dry paper meconium test to be the most useful and following to organize central screening center. Skin chloride system being fast and easy test is too expensive to be introduced as mass screening. Results of pilot screening study for congenital hypothyreosis point out the necessity for the mass diagnostic of this disorders. Choice of the test however is connected with economical aspects of the screening procedure.

[A new type of sweat test and its use in children]. / Nowy rodzaj testu potowego i jego zastosowanie u dzieci.

Chloride concentration in sweat was determined by the method of Skine Chloride System of Orion's Model 417 in 100 healthy children and 42 CF children. Statistically significant differences were demonstrated in chloride concentration in sweat between the group of healthy children and CF children. The results of our investigations suggest, that this method is useful in the diagnosis of CF in childhood.

[Determination of the ribonuclease activity of the duodenal contents of children as a test in the differential diagnosis of cystic fibrosis and pancreatic exocrine insufficiency]. / Oznaczanie aktywnosci rybonukleazy w tresci dwunastniczej u dzieci jako test róznicujacy cystic fibrosis od niewydolnosci zewna trzwydzielniczej trzustki.

Ribonuclease activity was determined in the duodenal contents of healthy children, in children with cystic fibrosis and with pancreatic exocrine insufficiency. Assays were made at 7.0, in 0.05 M buffer without and with 0.35 M NaCl. The ratio of activity in 0.35 M NaCl to that in buffer alone was found to be 2.4 +/- 1.3 for children with cystic fibrosis as compared with 12 +/- 5 for the control group and children with pancreatic exocrine insufficiency.