FERM protein EPB41L5 is a novel member of the mammalian CRB-MPP5 polarity complex

Cell polarity is induced and maintained by separation of the apical and basolateral domains through specialized cell-cell junctions. The Crumbs protein and its binding partners are involved in formation and stabilization of adherens junctions. In this study, we describe a novel component of the mammalian Crumbs complex, the FERM domain protein EPB41L5, which associates with the intracellular domains of all three Crumbs homologs through its FERM domain. Surprisingly, the same FERM domain is involved in binding to the HOOK domain of MPP5/PALS1, a previously identified interactor of Crumbs. Co-expression and co-localization studies suggested that in several epithelial derived tissues Epb4.1l5 interacts with at least one Crumbs homolog, and with Mpp5. Although at early embryonic stages Epb4.1l5 is found at the basolateral membrane compartment, in adult tissues it co-localizes at the apical domain with Crumbs proteins and Mpp5. Overexpression of Epb4.1l5 in polarized MDCK cells affects tightness of cell junctions and results in disorganization of the tight junction markers ZO-1 and PATJ. Our results emphasize the importance of a conserved Crumbs-MPP5-EPB41L5 polarity complex in mammals.     

Effect of PEEP on induced constriction is enhanced in decorin-deficient mice

Decorin (Dcn), a small leucine-rich proteoglycan, is present in the extracellular matrix of the airways and lung tissues, contributes to lung mechanical properties, and its deposition is altered in asthma. The effect of Dcn deficiency on airway parenchymal interdependence was examined during induced bronchoconstriction. Studies were performed in C57Bl/6 mice in which the Dcn gene was disrupted by targeted deletion (Dcn(-/-)) and in wild-type controls (Dcn(+/+)). Mice were mechanically ventilated, and respiratory system impedance was measured during in vivo ventilation at positive end-expiratory pressure (PEEP) = 2 and 10 cmH(2)0, before and after aerosol delivery of methacholine (MCh). Length vs. tension curves in isolated tracheal rings were measured in vitro. Dcn distribution in +/+ mice airways was characterized by immunofluorescence; differences in collagen structure in Dcn(+/+) and Dcn(-/-) mouse lungs was examined by electron microscopy. MCh caused similar increases in airway resistance (Raw) and tissue elastance (H) in Dcn(+/+) and Dcn(-/-) mice. During MCh-induced constriction, increasing PEEP caused a decrease in Raw that was greater in Dcn(-/-) mice and a decrease in H in Dcn(-/-) mice only. Tracheal ring compliance was greater in Dcn (-/-) mice. Imaging studies showed that Dcn was deposited primarily in the airway adventitial layer in Dcn(+/+) mice; in Dcn(-/-) mice, collagen had an irregular appearance, especially in the lung periphery. These results show that lack of Dcn alters the normal interaction between airways and lung parenchyma; in asthma, changes in Dcn could potentially contribute to abnormal airway physiology.     

Nicotinamide N-methyltransferase upregulation inversely correlates with lymph node metastasis in oral squamous cell carcinoma

We investigated expression levels of Nicotinamide N-Methyltransferase (NNMT), an enzyme involved in the biotransformation of many drugs and xenobiotic compounds, in oral squamous cell carcinoma (OSCC). Measurements were performed by semi-quantitative RT-PCR and quantitative real-time PCR in tumor and matched adjacent healthy tissue. Interestingly, NNMT was up-regulated in most of the favorable OSCCs, while no marked NNMT expression alterations between tumor and normal mucosa were detected in most of the unfavorable OSCCs. Western blot and immunohistochemical analyses also were performed and the relationship between tumor characteristics and NNMT levels in OSCC were studied to evaluate the effectiveness of NNMT as a prognostic marker in the squamous cell carcinoma of the oral cavity. In summary, the present study suggests that NNMT may have potential as a biomarker and a therapeutic target for OSCC.     

High-throughput liberation of water-soluble yeast content by irreversible electropermeation (HT-irEP)

The article describes a high-throughput method for the liberation of water-soluble cell contents by exploiting the phenomenon of irreversible membrane electropermeation (HT-irEP). The method is exemplified in recombinant proteins and plasmid liberation from yeast Saccharomyces cerevisiae on the detectable level. Obtained extracts are pure enough to be readily applied for further analytical analysis such as enzyme assay, PCR, and so on. From the same HT-irEP extract, one can measure activity of the target protein and perform amplification of the corresponding gene from the DNA vector by PCR for recombinant protein with intracellular expression. Therefore, the method is suitable for the high-throughput screening (HTS) of yeast libraries where extracellular expression of recombinant protein is problematic. The method can be easily automated and integrated into existing HTS systems.     

Human trisomy 21 fibroblasts rescue methotrexate toxic effect after treatment with 5-methyl-tetrahydrofolate and 5-formyl-tetrahydrofolate

Trisomy 21 causes Down syndrome (DS), the most common human genetic disorder and the leading genetic cause of intellectual disability. The alteration of one-carbon metabolism was described as the possible metabolic cause of the intellectual disability development in subjects with DS. One of the biochemical pathways involved in the one-carbon group transfer is the folate cycle. The cytotoxic drug methotrexate (MTX) is a folic acid (FA) analogue which inhibits the activity of dihydrofolate reductase enzyme involved in the one-carbon metabolic cycle. Trisomy 21 cells are more sensitive to the MTX effect than euploid cells, and in 1986 Jérôme Lejeune and Coll. demonstrated that MTX was twice as toxic in trisomy 21 lymphocytes than in control cells. In the present work, the rescue effect on MTX toxicity mediated by FA and some of its derivatives, tetrahydrofolate (THF), 5-formyl-THF, and 5-methyl-THF, in both normal and trisomy 21 skin fibroblast cells, was evaluated. A statistically significant rescue effect was obtained by 5-formyl-THF, 5-methyl-THF, and their combination, administered together with MTX. In conclusion, trisomy 21 fibroblast cell lines showed a good response to the rescue effects of 5-formyl-THF and 5-methyl-THF on the MTX toxicity almost as normal cell lines.     

Сlass II histone deacetylases in the post-stroke recovery period—expression,cellular, and subcellular localization—promising targets for neuroprotection

Histone deacetylases (HDAC) inhibitors can protect nerve cells after a stroke, but it is unclear which HDAC isoform is involved in this effect. We studied cellular and intracellular rearrangement of class II HDACs at late periods after photothrombotic infarct (PTI) in the mouse sensorimotor cortex in the tissue surrounding the ischemia core and in the corresponding region of the contralateral hemisphere. We observed a decrease in HDAC4 in cortical neurons and an increase in its nuclear translocation. HDAC6 expression in neurons was also increased. Moreover, HDAC6-positive cells had elevated apoptosis. Tubostatin A (Tub A)-induced decrease in the activity of HDAC6 restored acetylation of α-tubulin during the early poststroke recovery period and reduced apoptosis of nerve cells thus protecting the brain tissue. Selective inhibition of HDAC6 elevated expression of growth-associated protein-43 (GAP43), which remained high up to 14 days after stroke and promoted axogenesis and recovery from the PTI-induced neurological deficit. Selective HDAC6 inhibitor Tub A markedly reduced neuronal death and increased acetylation of α-tubulin and the level of GAP43. Thus, HDAC6 inhibition could be a promising strategy for modulation of brain recovery as it can increase the intensity and reduce the duration of reparation processes in the brain after stroke.     

Exploring the predictive performance of several temperature measurements on Neotropical dung beetle assemblages: Methodological implications

Basic characteristics of species assemblages are frequently related to temperature variables recorded at a coarse‐grained scale. In this study, 15 min instant‐measurements of environmental and soil temperatures were recorded during 1 year in six Atlantic Forest sites of southern Brazil, ranging from 250 to 1,630 m a.s.l. These measurements were used to examine the comparative explanatory capacity of several temperature variables in predicting species richness and total or specific variations of dung beetle abundance. The results suggest that temperature measurements obtained during the survey period have the highest explanatory capacity. Furthermore, average temperature values seem to have a relatively higher explanatory capacity than absolute minimum or maximum values reflecting extreme conditions. In general, there is no rule in selecting a temperature variable when the objective involves explaining the variation in species abundances. Both soil and air variables can have similar explanatory capacities. The present results should be considered when designing future ecological studies in Neotropical conditions.     

Skeletal evidence of tuberculosis in a modern identified human skeletal collection (Certosa cemetery,Bologna, Italy)

The diagnosis of tuberculosis (TB) in osteoarcheological series relies on the identification of osseous lesions caused by the disease. The study of identified skeletal collections provides the opportunity to investigate the distribution of skeletal lesions in relation to this disease. The aim of this study was to examine the skeletal evidence for TB in late adolescent and adult individuals from the identified human collection of the Certosa cemetery of Bologna (Italy, 19th–20th c.). The sample group consists of 244 individuals (138 males, 106 females) ranging from 17 to 88 years of age. The sample was divided into three groups on the basis of the recorded cause of death: TB (N = 64), pulmonary non‐TB (N = 29), and other diseases (N = 151). Skeletal lesions reported to be related to TB were analyzed. The vertebral lesions were classified into three types: enlarged foramina (EnF, vascular foramina with diameter of 3–5 mm), erosions (ER), and other foramina (OtF, cavities of various shapes > 3 mm). A CT scan analysis was also performed on vertebral bodies. Some lesions were seldom present in our sample (e.g., tuberculous arthritis). OtF (23.7%) and subperiosteal new bone formation on ribs (54.2%) are significantly more frequent in the TB group with respect to the other groups. The CT scan analysis showed that the vertebrae of individuals who have died of TB may have internal cavities in the absence of external lesions. These traits represent useful elements in the paleopathological diagnosis of TB. Am J Phys Anthropol 157:389–401, 2015. © 2015 Wiley Periodicals, Inc.