Body composition as an indicator of metabolic changes in mice obtained by in vitro fertilization.

To identify body systems subject to epigenetic transformation during in vitro fertilization (IVF), comparative morphological and functional studies were performed on sexually mature offspring of outbred CD1 mice, specific-pathogen-free (SPF), obtained by IVF (experiment) and natural conception (control). The studies included assessment of age-related changes in body weight and composition, energy intake and expenditure, and glucose homeostasis. To level the effects caused by the different number of newborns in the control and in the experiment, the size of the fed litters was halved in the control females. Males obtained using the IVF procedure were superior in body weight compared to control males in all age groups. As was shown by analysis of variance with experiment/control factors, gender, age (7, 10 and 20 weeks), the IVF procedure had a statistically significant and unidirectional effect on body composition. At the same time, IVF offspring outperformed control individuals in relative fat content, but were behind in terms of lean mass. The effect of the interaction of factors was not statistically significant. IVF offspring of both sexes had higher fat to lean mass ratios (FLR). Since adipose tissue contributes significantly less to total energy intake compared to muscle, the main component of lean mass, it is not surprising that at the same level of IVF locomotor activity offspring consumed less food than controls. When converted to one gram of body weight, this difference reached 19 %. One of the consequences of reduced utilization of IVF energy substrates by offspring is a decrease in their tolerance to glucose loading. The integral criterion for the effectiveness of restoring the initial glucose level is the area under the curve (AUC), the value of which was 2.5 (males) and 3.2 (females) times higher in IVF offspring compared to the corresponding control. Thus, the totality of our original and literature data shows an increase in the risk of metabolic disorders in IVF offspring, which is confirmed by epidemiological studies of a relatively young cohort of people born using assisted reproductive technologies.

Bioaccumulation as a method of removing psychoactive compounds from wastewater using aquatic plants.

Since WWTPs are not able to eliminate all psychoactive pharmaceuticals, these compounds become a part of the aquatic ecosystem. Our results indicate that compounds such as codeine or citalopram are eliminated with low efficiency (<38%), and compounds such as venlafaxine, oxazepam, or tramadol even with almost no efficiency. Lower elimination efficiency may be caused by the accumulation of these compounds in the wastewater treatment process. This study is focused on the possibility to remove problematic psychoactive compounds using aquatic plants. HPLC-MS analysis of the leaf extract obtained from studied plants showed that the amount of accumulated methamphetamine was highest in Pistia stratiotes and lower in the leaves of Limnophila sessiliflora and Cabomba caroliniana. However, tramadol and venlafaxine were accumulated considerably only in Cabomba caroliniana. Our study demonstrates that especially these three compounds - tramadol, venlafaxine, and methamphetamine, are accumulated in aquatic plants and can be removed from the aquatic environment. In our study was also observed that helophytic aquatic plants show a higher ability to remove psychoactive compounds from wastewater. Iris pseudacorus showed the best results in selected pharmaceuticals removal with no bioaccumulation effect in leaves or roots.

Ultrafast laser diode thermal desorption method for analysis of representative pharmaceuticals in soil leachate samples.

We developed and evaluated a novel analytical method combining ambient ionization technique - laser diode thermal desorption with chemical ionization (LDTD-APCI) and tandem mass spectrometry detection. The LDTD/APCI-MS/MS method was developed for determination of representative pharmaceuticals from different classes (carbamazepine, sulfamethoxazole, irbesartan, fexofenadine) in leachate samples from soil sorption experimentation. We then optimized laser pattern, laser energy and spiked sample volume, which are crucial parameters for this LDTD/APCI-MS/MS method. We further identified utility of a chelating agent (Na2-EDTA) to obtain the highest achievable and reproducible signal of target analytes. Achieved method performance parameters (LODs, LOQs, trueness and precision) were comparable with those obtained from LC-MS/MS. However, application of this novel LDTD/APCI-MS/MS method reduced analysis time by two orders of magnitude (to 12 s), compared to more conventional LC-MS/MS approaches, without use of organic solvents. We expect this novel method will reduce costs and increase throughput for future analyses of pharmaceuticals in the environment while advancing a timely principle of green chemistry.

Cells transformed by murine herpesvirus 68 (MHV-68) release compounds with transforming and transformed phenotype suppressing activity resembling growth factors.

In this study, we investigated the medium of three cell lines transformed with murine herpesvirus 68 (MHV-68) in vitro and in vivo, 68/HDF, 68/NIH3T3, and S11E, for the presence of compounds resembling growth factors of some herpesviruses which have displayed transforming and transformed phenotype suppressing activity in normal and tumor cells. When any of spent medium was added to cell culture we observed the onset of transformed phenotype in baby hamster kidney cells (BHK-21) cells and transformed phenotype suppressing activity in tumor human epithelial cells (HeLa). In media tested, we have identified the presence of putative growth factor related to MHV-68 (MHGF-68). Its bivalent properties have been blocked entirely by antisera against MHV-68 and two monoclonal antibodies against glycoprotein B (gB) of MHV-68 suggesting viral origin of MHGF-68. The results of initial efforts to separate MHGF-68 on FPLC Sephadex G15 column in the absence of salts revealed the loss of its transforming activity but transformed phenotype suppressing activity retained. On the other hand, the use of methanol-water mobile phase on RP-HPLC C18 column allowed separation of MHGF-68 to two compounds. Both separated fractions, had only the transforming activity to normal cells. Further experiments exploring the nature and the structure of hitherto unknown MHGF-68 are now in the progress to characterize its molecular and biological properties.

Clinical correlations of potential activity markers in systemic sclerosis.

The objectives of the project were the following: (1) to establish a group of patients with a confirmed diagnosis of systemic sclerosis (Ssc), (2) to perform a detailed entrance examination of each patient, (3) to determine concentrations of potential activity markers, and (4) to make a comprehensive examination of each patient 1 year after inclusion into the study. A total of 49 patients were examined, 36 with a limited form of SSc, 9 with diffuse SSc, and 4 with other forms of SSc. We determined plasma or serum levels of the N-terminal propeptide of procollagen type III (NPIIIP), interleukin-6 (IL-6), soluble receptor for interleukin-2 (sIL-2r), soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular adhesion molecule-1 (sVCAM-1), von Willebrand factor antigen (vWFAg), and big endothelin-1 (BET-1) using commercial kits, and urinary excretion of pyridinoline (PYR) and deoxypyridinoline (D-PYR) using high-performance liquid chromatography. Correlations of these markers with selected clinical data were calculated. The mean levels of all potential activity markers were increased compared with normal values, but differences were not significant. The levels of NPIIIP, D-PYR, and IL-6 were normal. The measured values after 1 year did not differ from the entry values. At entry, NPIIIP concentrations correlated with the finger-to-palm distance, and D-PYR corresponded with findings on a simplified health assessment questionnaire (FQ). IL-6 levels correlated with the leukocyte count, sIL-2r with the FQ, and ET-1 with the diffuse lung capacity for carbon monoxide. In general, we found only a few clinical correlates of potential activity markers. Our data confirmed the correlations of collagen metabolism markers with skin involvement and FQ, as was reported previously. Larger studies in this field are needed.