Postnatal exposure to di-(2-ethylhexyl)phthalate alters cardiac insulin signaling molecules and GLUT4Ser488 phosphorylation in male rat offspring

Di-(2-ethylhexyl)phthalate (DEHP), a distinctive endocrine-disrupting chemical, is widely used as a plasticizer in a variety of consumer products. It can easily cross the placenta and enter breast milk and then it is rapidly absorbed by offspring. Since it is generally accepted that individuals are more sensitive to chemical exposure during vital developmental periods, we investigated whether DEHP exposure during lactation affects cardiac insulin signaling and glucose homeostasis in the F₁ male rat offspring at postnatal day 22 (PND22). Lactating Wistar rats were administered with DEHP (1, 10, and 100 mg/kg/d) or olive oil from lactation day 1 to 21 by oral gavage. All the male pups were perfused and killed on PND22. On the day before the killing, they were kept for fasting overnight and blood was collected. The cardiac muscle was dissected out, washed in ice-cold physiological saline repeatedly and used for the assay of various parameters. DEHP-exposed offspring had significantly lower body weight than the control. DEHP-exposed offspring showed elevated blood glucose, decreased ¹⁴C-2-deoxyglucose uptake and ¹⁴C-glucose oxidation in cardiac muscle at PND22. The concentration of upstream insulin signaling molecules such as insulin receptor subunit β (InsRβ) and insulin receptor substrate 1 (IRS1) were downregulated in DEHP-exposed offspring. However, no significant alterations were observed in protein kinase B (Akt) and Akt substrate of 160 kDa (AS160). Surprisingly, phosphorylation of IRS1 ^Tyr632 and Akt ^Ser473 were diminished. Low levels of glucose transporter type 4 (GLUT4) protein and increased GLUT4 ^Ser488 phosphorylation which decreases its intrinsic activity and translocation towards plasma membrane were also recorded. Lactational DEHP exposure predisposes F ₁ male offspring to cardiac glucometabolic disorders at PND22, which may impair cardiac function.     

Interaction between Cymbidium aloifolium and Apis cerana: Incidence of an outlier in modular pollination network of oil flowers

So far, oil‐rewarding flowers are known to be pollinated only by oil‐collecting bees, which gather and use lipids for larval feed and nest building. As honeybees do not have oil‐collecting appendages on their legs, they have not been associated with pollination of such flowers. In a predominantly Apis pollinated and food deceptive clade of wild Cymbidiums, we investigated the reproductive strategy of Cymbidium aloifolium, hitherto unknown for its floral oil reward. Our study demonstrates the requisites for establishment of mutualistic interaction between the oil flower and Apis cerana indica, a corbiculate bee. Success in pollination requires learning by honeybees to access the food reward, thereby displaying cognitive ability of the pollinator to access the customized reward. Morphometric matching between orchid flowers and the pollinator, and that between pollinia and stigmatic cavity also appear to be essential in the pollination success. Absence of pollinator competition and prolonged flower‐handling time are suggested to promote floral constancy. The present study highlights the need to explore the spectrum of pollination rewards pursued by honeybees, which may include unconventional composition of floral resources.     

Galactomannanases Man2 and Man5 from Thermotoga species: growth physiology on galactomannans, gene sequence analysis, and biochemical properties of recombinant enzymes

The enzymatic hydrolysis of mannan-based hemicelluloses is technologically important for applications ranging from pulp and paper processing to food processing to gas and oil well stimulation. In many cases, thermostability and activity at elevated temperatures can be advantageous. To this end, the genes encoding beta-mannosidase (man2) and beta-mannanase (man5) from the hyperthermophilic bacteria Thermotoga neapolitana 5068 and Thermotoga maritima were isolated, cloned, and expressed in Escherichia coli. The amino acid sequences for the mannosidases from these organisms were 77% identical and corresponded to proteins with an M(r) of approximately 92 kDa. The translated nucleotide sequences for the beta-mannanase genes (man5) encoded polypeptides with an M(r) of 76 kDa that exhibited 84% amino acid sequence identity. The recombinant versions of Man2 and Man5 had similar respective biochemical and biophysical properties, which were also comparable to those determined for the native versions of these enzymes in T. neapolitana. The optimal temperature and pH for the recombinant Man2 and Man5 from both organisms were approximately 90 degrees C and 7.0, respectively. The presence of Man2 and Man5 in these two Thermotoga species indicates that galactomannan is a potential growth substrate. This was supported by the fact that beta-mannanase and beta-mannosidase activities were significantly stimulated when T. neapolitana was grown on guar or carob galactomannan. Maximum cell densities increased by at least tenfold when either guar or carob galactomannan was added to the growth medium. For T. neapolitana grown on guar at 83 degrees C, Man5 was secreted into the culture media, whereas Man2 was intracellular. These localizations were consistent with the presence and lack of signal peptides for Man5 and Man2, respectively. The identification of the galactomannan-degrading enzymes in these Thermotoga species adds to the list of biotechnologically important hemicellulases produced by members of this hyperthermophilic genera.     

The role of glutathione in cancer

Glutathione is an abundant natural tripeptide found within almost all cells. Glutathione is highly reactive and is often found conjugated to other molecules via its sulfhydryl moiety. It instils several vital roles within a cell including antioxidation, maintenance of the redox state, modulation of the immune response and detoxification of xenobiotics. With respect to cancer, glutathione metabolism is able to play both protective and pathogenic roles. It is crucial in the removal and detoxification of carcinogens, and alterations in this pathway, can have a profound effect on cell survival. However, by conferring resistance to a number of chemotherapeutic drugs, elevated levels of glutathione in tumour cells are able to protect such cells in bone marrow, breast, colon, larynx and lung cancers. Here we present a number of studies investigating the role of glutathione in promoting cancer, impeding chemotherapy, and the use of glutathione modulation to enhance anti-neoplastic therapy.     

Replication-mediated instability of the GAA triplet repeat mutation in Friedreich ataxia

Friedreich ataxia is caused by the expansion of a polymorphic and unstable GAA triplet repeat in the FRDA gene, but the mechanisms for its instability are poorly understood. Replication of (GAA•TTC)_n sequences (9–105 triplets) in plasmids propagated in Escherichia coli displayed length- and orientation-dependent instability. There were small length variations upon replication in both orientations, but large contractions were frequently observed when GAA was the lagging strand template. DNA replication was also significantly slower in this orientation. To evaluate the physiological relevance of our findings, we analyzed peripheral leukocytes from human subjects carrying repeats of similar length (8–107 triplets). Analysis of 9400 somatic FRDA molecules using small-pool PCR revealed a similar mutational spectrum, including large contractions. The threshold length for the initiation of somatic instability in vivo was between 40 and 44 triplets, corresponding to the length of a eukaryotic Okazaki fragment. Consistent with the stabilization of premutation alleles during germline transmission, we also found that instability of somatic cells in vivo and repeats propagated in E.coli were abrogated by (GAGGAA)_n hexanucleotide interruptions. Our data demonstrate that the GAA triplet repeat mutation in Friedreich ataxia is destabilized, frequently undergoing large contractions, during DNA replication.     

Fractal dimension in endometrial carcinoma

OBJECTIVE: To mathematically assess in a pilot study, endometrial glandular margin irregularity in simple hyperplasia, complex atypical hyperplasia and well-differentiated endometrial carcinoma with the help of box counting of fractal dimension and to discriminate these lesions on the basis of box counting of fractal dimension of the gland. STUDY DESIGN: Ten cases each of endometrial simple hyperplasia (without atypia), complex hyperplasia with atypia and endometrial carcinoma (well-differentiated, endometrioid) were assessed in the study. Five fields at 20 x magnification from each case were randomly selected, and the glands were outlined with the help of a pointer. Using the box counting method, the fractal dimension of each case was measured. RESULTS: Mean fractal dimension in simple hyperplasia, complex atypical hyperplasia and endometrial carcinoma was, 0.899 +/- 0.13, 0.932 +/- 0.042 and 0.939 +/- 0.02, respectively. Statistical analysis showed that the fractal dimension of glands of simple hyperplasia were significantly different from that of complex atypical hyperplasia and endometrial carcinoma (P = .041 and .013, respectively, ANOVA). However, there was no significant difference in fractal dimension between glands of complex hyperplasia and of endometrial carcinoma (P = .659, ANOVA). CONCLUSION: This study provides mathematical (objective) assessment of the measurement of glandular margin irregularities in simple hyperplasia, complex atypical hyperplasia and endometrial carcinoma. Fractal dimension of gland margin may have diagnostic potential in the future.     

Genetic admixture of European FRDA genes is the cause of Friedreich ataxia in the Mexican population

Friedreich ataxia accounts for approximately 75% of European recessive ataxia patients. Approximately 98% of pathogenic chromosomes have large expansions of a GAA triplet repeat in the FRDA gene (E alleles), and strong linkage disequilibrium among polymorphisms spanning the FRDA locus indicates a common origin for all European E alleles. In contrast, we found that only 14 of 151 (9.3%) Mexican Mestizo patients with recessive ataxia were homozygous for E alleles. Analysis of polymorphisms spanning the FRDA locus revealed that all Mestizo E alleles had the common European haplotype, indicating that they share a single origin. Genetic admixture levels were determined, which revealed that the relative contributions to the Mestizo FRDA gene pool by Native American and European genes were 76-87% and 13-24%, respectively, commensurate with the observed low prevalence of Friedreich ataxia in Mestizos. This indicates that Friedreich ataxia in Mexican Mestizos is due to genetic admixture of European mutant FRDA genes in the Native American gene pool that existed prior to contact with Europeans.