Elastic fiber assembly in the adult mouse pubic symphysis during pregnancy and postpartum

Impairment of pelvic organ support has been described in mice with genetic modifications of the proteins involved in elastogenesis, such as lysyl oxidase-like 1 (LOXL1) and fibulin 5. During pregnancy, elastic fiber-enriched pelvic tissues are modified to allow safe delivery. In addition, the mouse pubic symphysis is remodeled in a hormone-controlled process that entails the modification of the fibrocartilage into an interpubic ligament (IpL) and the relaxation of this ligament. After first parturition, recovery occurs to ensure pelvic tissue homeostasis. Because ligaments are the main supports of the pelvic organs, this study aimed to evaluate elastogenesis in the IpL during mouse pregnancy and postpartum. Accordingly, virgin, pregnant, and postpartum C57BL/6 mice were studied using light, confocal, and transmission electron microscopy as well as Western blots and real-time PCR. Female mice exhibited the separation of the pubic bones and the formation, relaxation, and postpartum recovery of the IpL. By the time the IpL was formed, the elastic fibers had increased in profile length and diameter, and they consisted of small conglomerates of amorphous material distributed among the bundles of microfibrils. Our analyses also indicated that elastin/tropoelastin, fibrillin 1, LOXL1/Loxl1, and fibulin 5 were spatially and temporally regulated, suggesting that these molecules may contribute to the synthesis of new elastic fibers during IpL development. Overall, this work revealed that adult elastogenesis may be important to assure the elasticity of the pelvic girdle during preparation for parturition and postpartum recovery. This finding may contribute to our understanding of pathological processes involving elastogenesis in the reproductive tract.     

Has Ombrëdanne's method of hypospadic urethra reconstruction been ignored with reason?

From January 1970 to December 1979 inclusive, 193 boys (aged 2 to 16) underwent surgery for distal hypospadia using Ombredanne's method at the Department of Pediatric Surgery University Hospital Center Rijeka and at the Department of Pediatric Surgery Zagreb. Follow-up period was 7 to 20 years (mean 13.4). 20 (10.36%) subjects had post-operative organic complications and 15 (7.77%) of them required surgical correction. According to these findings, the success rate using Ombredanne's method of reconstruction of the hypospadic urethra in no way lags behind the success rate using MAGPI and Mathieu's methods as well as Preputial island flap urethroplasty for analogous cases. Out of 193 subjects who underwent surgery, 80 (41.45%) of those who were sexually mature and had normal psychosexual development were questioned. In this sample, 75 (93.75%) were satisfied with the post-operative appearance of the penis while only 5 (6.25%) were dissatisfied, 3 of which had hypoplastic penis. In 78 (97.50%) subjects questioned, the post-operative urinary squirt was normal and two of them had weak urinary squirt (2.50%), due to meatal stenosis. In conclusion, Ombredanne's method of reconstruction of the urethra in boys with distal hypospadia is equally successful as other methods used for this purpose.     

Synthesis and anti-aromatase activity of some new steroidal D-lactones

Starting from D-seco derivatives of 5-androstene 1-3, the D-homo lactones, 4 and 5, were synthesized. By the Oppenauer oxidation and/or by dehydration of 4 and 5 with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) or 2,3,5,6-tetrachloro-1,4-benzoquinone (chloranil), the corresponding D-lactones 6-12 were obtained. The structures of 6 and 10 were unambiguously proved by the appropriate X-ray structural analysis. Anti-aromatase assay showed that tested compounds possess inhibition potency, however, two to four times smaller (IC50 from 0.2 to 0.7 microM, respectively) in comparison to aminoglutethimide (AG).     

The influence of 4-hydroxy-2-nonenal on proliferation, differentiation and apoptosis of human osteosarcoma cells

The product of lipid peroxidation, 4-hydroxy-2-nonenal (HNE) is known to cause cell death at high concentrations, while at lower concentrations it can influence cell proliferation and differentiation. In our experiments we used human osteosarcoma cells (HOS), to test the influence of HNE on cell proliferation, differentiation and induction of apoptosis. Apoptosis induction was estimated by TiterTACS TUNEL test. The cells were in parallel counted and the DAPI staining method was used to distinguish between apoptotic and necrotic cells as well as to define the proportion of cells in mitosis. To test the influence of HNE on HOS cell differentiation, cells were treated every second day with HNE. After 10 days, the cells were stained for alkaline phosphatase, a marker for osteoblast differentiation. Cell growth inhibition was caused by supraphysiological concentrations of 10 or 100 microM HNE, while apoptosis was induced with supraphysiological as well as by the physiological amount of the aldehyde (1 microM). Necrosis appeared when cells were treated with 10 or 100 microM, but not with 1 microM HNE. The proportion of cells in mitosis gradually declined with increased HNE concentration. Multiple exposures of HOS cells to 10 microM HNE prevented HOS cell differentiation. These results indicated that HNE inhibits proliferation and differentiation of HOS cells in the same concentration dependent manner as it causes apoptosis. We thus assume that HNE might be one of the important signaling molecules regulating the growth of the human osteosarcoma cells.     

Evidence for a natural hybrid of peacock bass (Cichla monoculus vs Cichla temensis) based on esterase electrophoretic patterns

Esterase (Est) and esterase-D (Est-D) electrophoretic patterns identified by starch gel electrophoresis of skeletal muscle protein extracts of 184 specimens of three species of peacock bass, locally known as tucunares (Cichla monoculus, C. temensis and Cichla sp), plus four specimens of a supposed hybrid (C. monoculus vs C. temensis), collected from the Central Amazon, were examined to determine if they could aid in identifying a supposed hybrid between C. monoculus and C. temensis. Six zones of electrophoretic activity were found with these enzyme systems. The Est enzyme showed one zone of activity, formed by bands 1, 2 and 3, plus three zones of activity, presumably controlled by Est-1, 2 and 3 loci. The Est-D enzyme had two zones of activity, presumably controlled by Est-D1 and Est-D2 loci. Cichla monoculus and C. temensis shared band 2 and alleles Est-1(1), Est-2(1), Est-3(2), and Est-D1(1), and therefore these were useless for identifying hybrids between the two species. However, a probable hybrid pattern of bands 1, 2, and 3, presumably generated by a combination of pattern 12 from C. monoculus with pattern 23 from C. temensis, resulting from a possible cross between these two species, was detected. Although the Est-D2 locus cannot be considered an ideal diagnostic marker for identifying the supposed hybrid (C. monoculus vs C. temensis), as it is polymorphic, it proved to be useful for determining the origin of the hybrid, i.e., which parental species were involved in the hybridization process.     

Allozyme Differentiation of Four Populations of Hypostomus (Teleostei: Loricariidae) from Ribeirã;o Keller, a Small Stream in the Upper Rio Paraná Basin, Brazil

Hypostomus hermanni (Ihering, 1905) and three other morphotypes of the genus Hypostomus were collected from the Ribeirã;o Keller, a small tributary of Rio Ivaí, a tributary of upper Rio Paraná. An allozyme analysis of 25 gene loci revealed diagnostic loci alleles for each population and there were fixed differences between some of them at some loci. Thus all populations were genetically distinct, although there were many common alleles. Heterozygosity (H _e) ranged from 0.059 in Hypostomus sp. 2 to 0.144 in Hypostomus sp. 1 and was higher than the average for other species of Hypostomus and also for fish in general. H. hermanni and Hypostomus sp. 1 were more similar to each other (I= 0.878) whereas H. hermanni and Hypostomus sp. 2 showed the least genetic identity (I= 0.392).     

Identifying anisotropic constraints in multiply labeled bacteriorhodopsin by 15N MAOSS NMR: a general approach to structural studies of membrane proteins

Structural models of membrane proteins can be refined with sets of multiple orientation constraints derived from structural NMR studies of specifically labeled amino acids. The magic angle oriented sample spinning (MAOSS) NMR approach was used to determine a set of orientational constraints in bacteriorhodopsin (bR) in the purple membrane (PM). This method combines the benefits of magic angle spinning (MAS), i.e., improved sensitivity and resolution, with the ability to measure the orientation of anisotropic interactions, which provide important structural information. The nine methionine residues in bacteriorhodopsin were isotopically (15)N labeled and spectra simplified by deuterium exchange before cross-polarization magic angle spinning (CPMAS) experiments. The orientation of the principal axes of the (15)N chemical shift anisotropy (CSA) tensors was determined with respect to the membrane normal for five of six residual resonances by analysis of relative spinning sideband intensities. The applicability of this approach to large proteins embedded in a membrane environment is discussed in light of these results.     

Molecular variants of human papilloma viruses type 16 and 6 in women with different cytological results detected by RFLP analysis

HPV infections are common and the presence of the same high-risk type in cervical specimens can be due to reinfection or persistence. Persistent infection is the most important predictor for development of cervical carcinoma. The aim of this study was to validate PCR-RFLP with two sets of primers: MY09/MY11 that amplify a fragment of L1 and P1/P2 that amplify a fragment of E1 ORF. PCR product of MY09/MY11 was digested with a set of 6 restriction enzymes (RE) and PCR product of P1/P2 with a set of 12 RE. Cervical samples from 110 women patients of the University Gynecologic Clinic CHC Zagreb were analyzed. There were 98 (89.1%) PCR positive samples detected with P1/P2 primers, and 94 (85.5%) PCR positive samples detected with MY09/MY11 primers. Seven HPV types were detected with P1/P2-RFLP technique and 17 with MY09/MY11-RFLP PCR positive samples amplified with both primer pairs agreed with each other in 82 samples; 16 samples were only positive with P1/P2 and 12 samples were only positive by MY09/MY11. HPV 16 was detected in 39 samples with MY09/11-RFLP, out of these two variants (two different patterns) were found with P1/P2 using Dde I, Hae III and Eco I. HPV 6 was detected in 9 samples with MY09/11-RFLP, out of these two variants were found with P1/P2 using HinfI. Combining these two PCR-RFLP methods subtypes of HPV 16 and HPV 6 were detected.     

Environmental factors influence on chemical polymorphism of the essential oils of Lychnophora ericoides

Lychnophora ericoides is a Brazilian medicinal plant used in folk medicine as an anti-inflammatory and analgesic agent. The essential oils from leaves of two populations with and without scent, collected at 2-month intervals during an 1-year period, were analysed by GC-MS. The results were submitted to principal component and cluster analysis which allowed two groups of essential oils to be distinguished with respect to sampling site and scent: cluster I (Vianópolis site, with specimens exhibiting an aromatic scent) containing a high percentage of alpha-bisabolol (44.7-76.4%) and alpha-cadinol (10.9-23.5%), and cluster II (Cristalina site, with specimens without scent) characterised by a high content of (E)-nerolidol (31.3-47.1%) and ar-dihydro-turmerone (4.8-15.4%). The canonical discriminant analysis showed that using the data set of the seven sampling months and (E)-nerolidol and alpha-bisabolol as predictable variables, it was possible to distinguish between the samples harvested according to Cerrado seasons, dry winter (May-September) and humid summer (November-March). In addition, canonical correlation analysis between the soil sampling sites and the populations revealed a significant relationship between oil components and edaphic factors. Oxygenated sesquiterpenes and potential acidity, Al saturation, cationic exchange capacity, silt, and sand load as the first canonical variate were fairly strongly related to samples collected in Vianópolis site. On the other hand, monoterpenes and sesquiterpene hydrocarbons were strongly related to chemical balance in soils (organic matter, P and base saturation), which is related to samples at the Cristalina site. The chemovariation observed appears to be environmentally determined.     

Circulating gastrin is increased in hemochromatosis

Gastric acid production is important in intestinal iron absorption. The peptide hormone gastrin exists in both amidated and non-amidated forms, which stimulate and potentiate gastric acid secretion, respectively. Since non-amidated gastrins require ferric ions for biological activity in vitro, this study investigated the connection between iron status and gastrin by measurement of circulating gastrin concentrations in mice and humans with hemochromatosis. Gastrin concentrations are increased in the plasma and gastric mucosa of Hfe(-/-) mice, and in the sera of humans with HFE-related hemochromatosis. The discovery of a relationship between iron status and circulating gastrin concentrations opens a new perspective on the mechanisms of iron homeostasis.