Interaction between IRF6 and TGFA Genes Contribute to the Risk of Nonsyndromic Cleft Lip/Palate

Previous evidence from tooth agenesis studies suggested IRF6 and TGFA interact. Since tooth agenesis is commonly found in individuals with cleft lip/palate (CL/P), we used four large cohorts to evaluate if IRF6 and TGFA interaction contributes to CL/P. Markers within and flanking IRF6 and TGFA genes were tested using Taqman or SYBR green chemistries for case-control analyses in 1,000 Brazilian individuals. We looked for evidence of gene-gene interaction between IRF6 and TGFA by testing if markers associated with CL/P were overtransmitted together in the case-control Brazilian dataset and in the additional family datasets. Genotypes for an additional 142 case-parent trios from South America drawn from the Latin American Collaborative Study of Congenital Malformations (ECLAMC), 154 cases from Latvia, and 8,717 individuals from several cohorts were available for replication of tests for interaction. Tgfa and Irf6 expression at critical stages during palatogenesis was analyzed in wild type and Irf6 knockout mice. Markers in and near IRF6 and TGFA were associated with CL/P in the Brazilian cohort (p<10⁻⁶). IRF6 was also associated with cleft palate (CP) with impaction of permanent teeth (p<10⁻⁶). Statistical evidence of interaction between IRF6 and TGFA was found in all data sets (p = 0.013 for Brazilians; p = 0.046 for ECLAMC; p = 10⁻⁶ for Latvians, and p = 0.003 for the 8,717 individuals). Tgfa was not expressed in the palatal tissues of Irf6 knockout mice. IRF6 and TGFA contribute to subsets of CL/P with specific dental anomalies. Moreover, this potential IRF6-TGFA interaction may account for as much as 1% to 10% of CL/P cases. The Irf6-knockout model further supports the evidence of IRF6-TGFA interaction found in humans.     

Dye coupling and connexin expression by cortical radial glia in the early postnatal subventricular zone

In this study, we have analyzed the specific contribution of the cortical radial glia (RG) for gap junctional communication (GJC) within the postnatal subventricular zone (SVZ). To specifically target RG as source of dye‐coupling in situ, we have developed a new technique that involves direct cell loading through the processes that reach the pial surface, with a mix of gap junction permeant (Lucifer yellow, LY) and nonpermeant (rhodamine‐conjugated dextran 3 KDa, RD) fluorochromes, the latter used as a marker for direct loaded cells. Tissue sections were analyzed for identification of directly loaded (LY+RD+) and coupled cells (LY+RD–) in the SVZ. Directly loaded cells were restricted to the region underlying the pial loading surface area. Coupled cells were distributed in a bistratified manner, along the outer dorsal surface of the SVZ and aligning the ventricle, leaving the SVZ core relatively free. Blocking GJC prior to pial loading greatly reduced dye coupling. Phenotypic analysis indicated that coupling by RG excludes neuroblasts and is mostly restricted to cells of glial lineage. Notwithstanding, no corresponding restriction to specific cell phenotype was found for two connexin isotypes, Cx43 and Cx45, in the postnatal SVZ. The extensive homocellular cell coupling by RG suggests an important role in the regulation of neurogenesis and functional compartmentalization of the postnatal SVZ. © 2012 Wiley Periodicals, Inc. Develop Neurobiol 2012     

EFFECTS OF HUMAN ACTIVITIES ON STRUCTURE AND COMPOSITION OF WOODY SPECIES OF THE NOKREK BIOSPHERE RESERVE OF MEGHALAYA,NORTHEAST INDIA

Aims Our study was conducted in the Nokrek Biosphere Reserve (NBR) in the Garo hills districts of Meghalaya, Northeast India. Our aim was to assess the effects of human activities on plant diversity,population structure and regeneration.Methods We selected a representative 1.2 hm2 stand in both the core and buffer zones of NBR. Structure and composition were determined by randomly sampling square quadrats, population structure was assessed by determining age structure, and regeneration was assessed by measuring densities of seedling, sapling and adult trees.Important findings More woody species were recorded from the core zone than the buffer zone (87 vs. 81 species), and there were a large number of tropical, temperate, and Sino-Himalayan, Burma-Malaysian and Malayan elements, primitive families and primitive genera. The trees were distributed in three distinct strata,canopy, subcanopy and sapling. Subcanopy and sapling layers had the highest species richness (81％ -88％ ). Lauraceae and Euphorbiaceae were the dominant families in terms of the number of species, and a large number of families were represented by single species. Most woody species (57 ％ - 79 ％ ) were contagiously distributed and had low frequency ( ＜ 20％ ). Although stand density was high in the buffer zone, its basal area was low compared to the stand in the core zone. Low similarity and high β-diversity indicate marked differences in species composition of the stands. Shannon diversity index was high in both the stands, while Simpson dominance index was low. The diameter-class distribution for dominant species revealed that the most had a large number of young individuals in their populations. Preponderance of tree seedlings, followed by a steep decline in population density of saplings and adult trees, indicated that the seedling to sapling stage was the most critical in the life cycle of the tree populations. Most species (42 ％ - 48 ％ ) had no regeneration,25 ％ - 35 ％ had good/fair regeneration, and the rest had poor regeneration or reoccurred as immigrants.     

Capillariid nematodes in Brazilian turkeys, Meleagris gallopavo (Galliformes, Phasianidae): pathology induced by Baruscapillaria obsignata and Eucoleus annulatus (Trichinelloidea, Capillariidae)

The pathology induced in turkeys (Meleagris gallopavo) by two capillariid nematodes, Baruscapillaria obsignata and Eucoleus annulatus is described together with data on prevalences, mean infection and range of worm burdens. B. obsignata occurred with a prevalence of 72.5% in the 40 examined hosts in a range of 2-461 nematodes and a mean intensity of 68.6, whereas E. annulatus was present in 2.5% of the animals, with a total amount of five recovered parasites. Gross lesions were not observed in the parasitized birds. Lesions due to B. obsignata mainly consisted of the thickening of intestinal villi with a mild mixed inflammatory infiltrate with the presence of mononuclear cells and heterophils. The lesions induced by E. annulatus were represented by foci of inflammatory infiltrate with heterophils in the crop epithelium and esophagus of a single infected female. These are the first pathological findings related to the presence of capillariid worms in turkeys to be reported in Brazil so far. Capillaria anatis, although present, was not pathogenic to the investigated turkeys.     

Simultaneous determination of sulfamethoxazole and trimethoprim in biological fluids for high-throughput analysis: comparison of HPLC with ultraviolet and tandem mass spectrometric detection

The comparison of two methods based on online solid phase extraction-liquid chromatography with UV (SPE-LC-UV) or mass spectrometry detection (SPE-LC-MS/MS) for the simultaneous quantification of sulfamethoxazole (SMZ) and trimethoprim (TMP) is presented. The methods were validated and proved to be accurate. The analysis of standard samples for SMZ at concentrations of 0.5, 1.5, 25 and 50microg/mL demonstrated a relative standard deviation of less than 6% for both methods (n=18), while TMP samples at concentrations of 0.05, 0.15, 1.5 and 5.0microg/mL were analyzed with R.S.D. of less than 4% (n=18). The method with mass spectrometric detection was approximately six times more sensitive than the method with ultraviolet detection. The total run time for the SPE-LC-MS/MS was 2.5min per sample as opposed to 18.0min for the SPE-LC-UV method. The method with MS detection in comparison with UV detection proved to be more rugged and was successfully applied to pharmacokinetics studies.     

Applying near-infrared spectroscopy in downstream processing: one calibration for multiple clarification processes of fermentation media

The use of near-infrared spectroscopy (NIRS) is demonstrated in the first downstream processing (DSP) steps of an active pharmaceutical ingredient (API) manufacturing process. The first method developed was designed to assess the API content in the filtrate stream (aqueous) of a rotary drum vacuum filter. The PLS method, built after spectral preprocessing and variable selection, had an accuracy of 0.01% (w/w) for an API operational range between 0.20 and 0.45% (w/w). The robustness and extrapolation ability of the calibration was proved when samples from ultrafiltration and nanofiltration processes, ranging from 0 to 2% (w/w), were linearly predicted (R2=0.99). The development of a robust calibration model is generally a very time-consuming task, and once established it is imperative that it can be useful for a long period of time. This work demonstrates that NIR procedures, when carefully developed, can be used in different process conditions and even in different process steps of similar unit operations.