Role of the Cldn6 cytoplasmic tail domain in membrane targeting and epidermal differentiation in vivo

It is widely recognized that the claudin (Cldn) family of four tetraspan transmembrane proteins is crucial for tight junction assembly and permeability barrier function; however, the precise role of the tail and loop domains in Cldn function is not understood. We hypothesized that the cytoplasmic tail domain of Cldn6 is crucial for membrane targeting and hence epidermal permeability barrier (EPB) formation. To test this hypothesis via a structure-function approach, we generated a tail deletion of Cldn6 (CDelta187) and evaluated its role in epidermal differentiation and EPB formation through its forced expression via the involucrin (Inv) promoter in the suprabasal compartment of the transgenic mouse epidermis. Even though a functional barrier formed, Inv-CDelta187 mice displayed histological and biochemical abnormalities in the epidermal differentiation program and stimulation of epidermal cell proliferation in both the basal and suprabasal compartments of the interfolliclar epidermis, leading to a thickening of the epidermis after 1 week of age that persisted throughout life. Although some membrane localization was evident, our studies also revealed a significant amount of not only Cldn6 but also Cldn10, Cldn11, and Cldn18 in the cytoplasm of transgenic epidermal cells as well as the activation of a protein-unfolding pathway. These findings demonstrate that the overexpression of a tail truncation mutant of Cldn6 mislocalizes Cldn6 and other Cldn proteins to the cytoplasm and triggers a postnatal increase in proliferation and aberrant differentiation of the epidermis, emphasizing the importance of the Cldn tail domain in membrane targeting and function in vivo.     

Chromosomal studies of male infertility

Prometaphase and metaphase chromosome analyses performed on 70 consecutive men with primary infertility (for a period of at least 2 years) revealed 8 (11.42%) men with some kind of chromosomal abnormality. The highest frequency of abnormal karyotypes (10%) was found among patients with azpospennia and the most frequent anomaly was 47, XXY chromosomal constitution, found in 6 (8.57%) patients. All the chromosomal aberrations found in this study was sex chromosomal type and we did not find any autosomal aberration. All patients with numerical chromosomal anomalies had azoospermia. The incidence of structural aberration in our study was 1.42%. Fifteen patients had different chromosomal variants (21.38%). We suggest that men with azoospermia should be considered for cytogenetic investigation and we report that "variants of the Y chromosome" have no influence on the sperm count (million/ml) and fertility of men.     

Culture of Helicobacter pylori from stool samples in children

We evaluated two protocols for isolation of Helicobacter pylori in stool from biopsied and nonbiopsied children. Twenty-three child patients whose presumptive positivity or negativity was diagnosed by endoscopy and a rapid urease test at site were used to compare biopsy-based tests with stool-based tests (H. pylori stool antigen test and stool culture). Their gastric activity and bacterial density were graded by the updated Sydney system. Biopsy and stool specimens were cultured on Campy-blood and Belo horizonte agar plates after enrichment in selective Campy-Thio medium. To compare two stool culture protocols, stools from 20 nonbiopsied children were tested by the HpSA test and cultured either as above or after treatment with cholestyramine. Grown colonies were screened by Gram staining, slide agglutination using anti-H. pylori monoclonal IgG; positive isolates were tested by biochemical tests and polymerase chain reaction for H. pylori-specific ureA gene. Coccoid H. pylori was isolated in stool samples from the biopsied patients whose bacterial density was two to four in histology. Their oxidase was slightly positive but became positive after two subcultures, while additional biochemical tests confirmed the isolation of H. pylori. Similar coccoid but oxidase positive H. pylori was isolated from three nonbiopsied children with the protocol of cholestyramine treatment only. The density of bacteria in the stomach may influence the recovery of H. pylori from stool; inactivation of bile with cholestyramine improves the yield in culture and favors isolation of an enhanced metabolic form of bacteria.     

Understanding Bactrocera dorsalis trapping to calibrate area-wide management

Knowing the dispersal of released insects and estimating the size of field populations are keys to the successful implementation of area-wide management (AWM) programmes based on the sterile insect technique (SIT), as they determine the release strategy of sterile males. Mark–release–recapture (MRR) is a common method used to estimate field populations and spatiotemporal dynamics. However, the extent to which the pest is attracted to lures is often difficult to identify, thereby biasing extrapolation to movement patterns and population size. We performed MRR experiments on the Oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), in a fruit-growing area in Senegal. Methyl eugenol and protein baits were used to trap males and females, respectively. After studying the suitability of marking B. dorsalis with fluorescent pigments at the laboratory, two releases of marked sterile flies were organized in the centre of an organic mango orchard, first in the late mango fruiting stage and second in the fruit setting stage. Traps were placed symmetrically along a 250 and 500 m radius from the release point. A very small fraction of the released individuals was recaptured: 0.45% in the first release and 3.08% in the second. Trapping of both sterile and wild flies was completely anisotropic. Sterile flies were mostly trapped at a relatively short distance (250 m) from the release point, in the first two days. Male trapping using methyl eugenol was highly effective, whereas the response of females to food bait traps was low. The direction of the wind was the main driver of recapture, with flies heading upwind. The results underline the importance of taking the odour plume around the traps into account when estimating populations, and the heterogeneous spread of the wild population in the landscape for the set-up of the release strategy of sterile insects for SIT-based AWM.     

Nested cladistic analysis of Brazilian populations of Drosophila serido.

The effects of Quaternary climatic cycles were investigated in Drosophila serido, a Brazilian cactophilic fly widely distributed outside the Amazonian region. Previous studies have indicated this species displays remarkable karyotypic, male genitalia, and mtDNA variation, so much so that it has been described as a species complex, or superspecies. In the present study we expand the analysis of the mtDNA COI gene on D. serido populations, particularly in central Brazil, by obtaining DNA sequences from 248 individuals distributed across 47 localities. This allowed us to perform a nested clade analysis to discriminate historical from recurrent forces shaping the evolution of D. serido populations. The nested analysis indicates one event of past fragmentation separating populations from south and central Brazil (referred to as type B) from populations in central and northeast Brazil (type D) and 15 other significant events. The most common outcome of our analysis was contiguous range expansion and we discuss why this was expected in D. serido. Our data indicate that D. serido has been distributed across Brazil at least since the Mid-Pleistocene, which contradicts the hypothesis of current distribution being determined by last glaciation cycle. Nonetheless, we present evidence that climatic cycles during the Quaternary and before have had a significant impact on the differentiation of D. serido in Brazil. Our study confirms the usefulness of the nested clade analysis for disentangling the effects of historical and present-day forces shaping the evolution and distribution of a taxon.     

A mobility-based cluster formation algorithm for wireless mobile ad-hoc networks

In the last decade, numerous efforts have been devoted to design efficient algorithms for clustering the wireless mobile ad-hoc networks (MANET) considering the network mobility characteristics. However, in existing algorithms, it is assumed that the mobility parameters of the networks are fixed, while they are stochastic and vary with time indeed. Therefore, the proposed clustering algorithms do not scale well in realistic MANETs, where the mobility parameters of the hosts freely and randomly change at any time. Finding the optimal solution to the cluster formation problem is incredibly difficult, if we assume that the movement direction and mobility speed of the hosts are random variables. This becomes harder when the probability distribution function of these random variables is assumed to be unknown. In this paper, we propose a learning automata-based weighted cluster formation algorithm called MCFA in which the mobility parameters of the hosts are assumed to be random variables with unknown distributions. In the proposed clustering algorithm, the expected relative mobility of each host with respect to all its neighbors is estimated by sampling its mobility parameters in various epochs. MCFA is a fully distributed algorithm in which each mobile independently chooses the neighboring host with the minimum expected relative mobility as its cluster-head. This is done based solely on the local information each host receives from its neighbors and the hosts need not to be synchronized. The experimental results show the superiority of MCFA over the best existing mobility-based clustering algorithms in terms of the number of clusters, cluster lifetime, reaffiliation rate, and control message overhead.     

Lactic acid production by loofah-immobilized Rhizopus oryzae through one-step fermentation process using starch substrate

Rhizopus oryzae PTCC 5263 capacity in synthesis of lactic acid (LA) from 10 g/l of soluble potato starch was determined using one-step fermentation process. Pellets were the favorable growing form of the free cells. The extent of the natural ability of the test fungus on biofilm formation on loofah sponge was examined by immobilizing R. oryzae (LIRO). The maximum LA concentration for the free cells and LIRO within 96 h was 3 and 4 g/l, respectively. In terms of specific starch utilization rate (\(q_{\text{s}}\)) and specific LA formation (\(q_{\text{p}}\)), LIRO performed more favorably compared to the free cells (\(q_{{{\text{s}}_{\text{F}} }} > q_{{{\text{s}}_{\text{LIRO}} }}\) and \(q_{{{\text{p}}_{\text{F}} }} < q_{{{\text{p}}_{\text{LIRO}} }}\)). Cell immobilization strategy was undertaken for the column reactor studies based on the statistically optimized levels of the inoculum size and temperature. Maximum production of the LA by the LIRO using an airlift reactor with net draft tube was 5 g/l obtainable within 48 h.

     

iPSC modeling of stage-specific leukemogenesis reveals BAALC as a key oncogene in severe congenital neutropenia

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