Endocrine regulation of the fasting response by PPARalpha-mediated induction of fibroblast growth factor 21

Peroxisome proliferator-activated receptor α (PPARα) regulates the utilization of fat as an energy source during starvation and is the molecular target for the fibrate dyslipidemia drugs. Here, we identify the endocrine hormone fibroblast growth factor 21 (FGF21) as a mediator of the pleiotropic actions of PPARα. FGF21 is induced directly by PPARα in liver in response to fasting and PPARα agonists. FGF21 in turn stimulates lipolysis in white adipose tissue and ketogenesis in liver. FGF21 also reduces physical activity and promotes torpor, a short-term hibernation-like state of regulated hypothermia that conserves energy. These findings demonstrate an unexpected role for the PPARα-FGF21 endocrine signaling pathway in regulating diverse metabolic and behavioral aspects of the adaptive response to starvation.     

Myeloid differentiation factor 88-independent Toll-like receptor pathway: Sustaining inflammation or promoting tolerance?

Toll-like receptor signaling represents a principal molecular pathway for host innate immunity. Mechanistically, it can be segregated into two distinct cascades: the myeloid differentiation factor 88-dependent and myeloid differentiation factor 88-independent (or Toll receptor-associated activator of interferon-mediated) cascades. Myeloid differentiation factor 88-dependent signaling is common to all the Toll-like receptors, except Toll-like receptor 3, which exclusively utilizes the myeloid differentiation factor 88-independent pathway. Based on recent evidences, a possible role for myeloid differentiation factor 88-independent pathway not only in sustaining inflammatory responses during Gram-negative infection but also in protective responses like endotoxin tolerance is proposed in this review. We hypothesize that the delayed kinetics of activation of the myeloid differentiation factor 88-independent pathway might be functionally aimed at tuning-down inflammatory reactions through promotion of cellular tolerization and possibly protecting hosts from inflammation-induced injury. Future studies will be needed to experimentally validate this idea and the crucial relevance of the myeloid differentiation factor 88-independent pathway.     

Isolated colostomy site recurrence in rectal cancer-two cases with review of literature

Background

Colostomy site carcinomas are rare with only eight cases reported in the world literature. Various etiological factors like adenoma-cancer sequence, bile acids, recurrent and persistent physical damage at the colostomy site by faecal matter due to associated stomal stenosis have been considered responsible. Two such cases are being reported and in both cases there was no evidence of any local recurrence in the pelvis or liver and distant metastasis. Both patients had received adjuvant chemotherapy following surgery.

Case presentation

First case was a 30-year-old male that had reported with large bowel obstruction due to an obstructing ulcero-proliferative growth (poorly differentiated adenocarcinoma) at the colostomy site after abdomino-perineal resection, performed for low rectal cancer six years previously. Wide local excision with microscopically free margins was performed with a satisfactory outcome. Four years later he presented with massive malignant ascites, cachexia and multiple liver metastasis and succumbed to his disease.Second case was a 47-year-old male that presented with acute large bowel obstruction due to an annular growth (well differentiated adenocarcinoma) in the upper rectum. He was managed by Hartmann's operation and the sigmoid colostomy was closed six months later. Five years following closure of colostomy, he presented with two parietal masses at the previous colostomy site scar, which, on fine needle aspiration cytology were found to be well-differentiated adenocarcinomas of colorectal type. Surgery in the form of wide local resection with free margins was performed. He presented again after five years with recurrence along the previous surgery scar and an incisional hernia and was managed by wide local excision along with hernioplasty. Follow-up of nine years following first surgery is satisfactory.

Conclusion

Colostomy site/scar recurrence of rectal carcinoma is rare and could be due to various etiological factors, although the exact causative mechanism is not known. Surgery with microscopically free margins is recommended in the absence of metastatic disease. Stenosis of the stoma is considered as one of the most important contributory factors and should be followed carefully.

     

A Method to Constrain Genome-Scale Models with 13C Labeling Data

Current limitations in quantitatively predicting biological behavior hinder our efforts to engineer biological systems to produce biofuels and other desired chemicals. Here, we present a new method for calculating metabolic fluxes, key targets in metabolic engineering, that incorporates data from ¹³C labeling experiments and genome-scale models. The data from ¹³C labeling experiments provide strong flux constraints that eliminate the need to assume an evolutionary optimization principle such as the growth rate optimization assumption used in Flux Balance Analysis (FBA). This effective constraining is achieved by making the simple but biologically relevant assumption that flux flows from core to peripheral metabolism and does not flow back. The new method is significantly more robust than FBA with respect to errors in genome-scale model reconstruction. Furthermore, it can provide a comprehensive picture of metabolite balancing and predictions for unmeasured extracellular fluxes as constrained by ¹³C labeling data. A comparison shows that the results of this new method are similar to those found through ¹³C Metabolic Flux Analysis (¹³C MFA) for central carbon metabolism but, additionally, it provides flux estimates for peripheral metabolism. The extra validation gained by matching 48 relative labeling measurements is used to identify where and why several existing COnstraint Based Reconstruction and Analysis (COBRA) flux prediction algorithms fail. We demonstrate how to use this knowledge to refine these methods and improve their predictive capabilities. This method provides a reliable base upon which to improve the design of biological systems.     

Molecular Evolution of Translin Superfamily Proteins Within the Genomes of Eubacteria, Archaea and Eukaryotes

Translin and its interacting partner protein, TRAX, are members of the translin superfamily. These proteins are involved in mRNA regulation and in promoting RISC activity by removing siRNA passenger strand cleavage products, and have been proposed to play roles in DNA repair and recombination. Both homomeric translin and heteromeric translin-TRAX complex bind to ssDNA and RNA; however, the heteromeric complex is a key activator in siRNA-mediated silencing in human and drosophila. The residues critical for RNase activity of the complex reside in TRAX sequence. Both translin and TRAX are well conserved in eukaryotes. In present work, a single translin superfamily protein is detected in Chloroflexi eubacteria, in the known phyla of archaea and in some unicellular eukaryotes. The prokaryotic proteins essentially share unique sequence motifs with eukaryotic TRAX, while the proteins possessing both the unique sequences and conserved indels of TRAX or translin can be identified from protists. Intriguingly, TRAX protein in all the known genomes of extant Chloroflexi share high sequence similarity and conserved indels with the archaeal protein, suggesting occurrence of TRAX at least at the time of Chloroflexi divergence as well as evolutionary relationship between Chloroflexi and archaea. The mirror phylogeny in phylogenetic tree, constructed using diverse translin and TRAX sequences, indicates gene duplication event leading to evolution of translin in unicellular eukaryotes, prior to divergence of multicellular eukayrotes. Since Chloroflexi has been debated to be near the last universal common ancestor, the present analysis indicates that TRAX may be useful to understand the tree of life.     

BRCA2 protein deficiency exaggerates doxorubicin-induced cardiomyocyte apoptosis and cardiac failure

The tumor suppressor breast cancer susceptibility gene 2 (BRCA2) plays an important role in the repair of DNA damage, and loss of BRCA2 predisposes carriers to breast and ovarian cancers. Doxorubicin (DOX) remains the cornerstone of chemotherapy in such individuals. However, it is often associated with cardiac failure, which once manifests carries a poor prognosis. Because BRCA2 regulates genome-wide stability and facilitates DNA damage repair, we hypothesized that loss of BRCA2 may increase susceptibility to DOX-induced cardiac failure. To this aim, we generated cardiomyocyte-specific BRCA2 knock-out (CM-BRCA2(-/-)) mice using the Cre-loxP technology and evaluated their basal and post-DOX treatment phenotypes. Although CM-BRCA2(-/-) mice exhibited no basal cardiac phenotype, DOX treatment resulted in markedly greater cardiac dysfunction and mortality in CM-BRCA2(-/-) mice compared with control mice. Apoptosis in left ventricular (LV) sections from CM-BRCA2(-/-) mice compared with that in corresponding sections from wild-type (WT) littermate controls was also significantly enhanced after DOX treatment. Microscopic examination of LV sections from DOX-treated CM-BRCA2(-/-) mice revealed a greater number of DNA double-stranded breaks and the absence of RAD51 focus formation, an essential marker of double-stranded break repair. The levels of p53 and the p53-related proapoptotic proteins p53-up-regulated modulator of apoptosis (PUMA) and Bax were significantly increased in samples from CM-BRCA2(-/-) mice. This corresponded with increased Bax to Bcl-2 ratios and elevated cytochrome c release in the LV sections of DOX-treated CM-BRCA2(-/-) mice. Taken together, these data suggest a critical and previously unrecognized role of BRCA2 as a gatekeeper of DOX-induced cardiomyocyte apoptosis and susceptibility to overt cardiac failure. Pharmacogenomic studies evaluating cardiac function in BRCA2 mutation carriers treated with doxorubicin are encouraged.     

CuO thin film based uric acid biosensor with enhanced response characteristics

An electrochemical approach for detection of individual single nucleotide polymorphisms (SNPs) based on nucleobase-conjugated apoferritin probe loaded with metal phosphate nanoparticles is reported. Coupling of the nucleotide-modified nanoparticle probe to the mutant sites of duplex DNA was induced by DNA polymerase I (Klenow fragment) to preserve Watson-Crick base-pairing rules. After sequential liquid hybridization of biotinylated DNA probes with mutant DNA and complementary DNA, the resulting duplex DNA helixes were captured to the surface of magnetic beads through a well known and specific biotin-streptavidin affinity binding. For signaling each of eight possible Single-nucleotide polymorphisms (SNPs), Pb, Cu, Cd and Zn phosphate-loaded apoferritin nanoparticle probes were linked to adenosine (A), cytidine (C), guanosine (G), and thymidine (T) mononucleotides, respectively. Monobase-conjugated apoferritin probes were coupled to the mutant sites of the formed duplex DNA in the presence of DNA polymerase. Electrochemical stripping analyses of the metals loaded in apoferritin nanoparticle probes provide a means for detection and quantification of mutant DNA. Each mutation captures different nucleotide-conjugated apoferritin probe and provide a distinct four-potential voltammogram, whose peak potentials reflect the identity of the mismatch. The method is sensitive enough to accurately determine AG mutation, as the most thermodynamically stable mismatch to detect, in the range of 50-600 pM. The proposed protocol provides a simple, fast, cost-effective, accurate and sensitive method for detection of SNPs.     

Synthesis and SAR of pyridothiazole substituted pyrimidine derived HCV replication inhibitors

Introduction of a nitrogen atom into the benzene ring of a previously identified HCV replication (replicase) benzothiazole inhibitor 1, resulted in the discovery of the more potent pyridothiazole analogues 3. The potency and PK properties of the compounds were attenuated by the introductions of various functionalities at the R(1), R(2) or R(3) positions of the molecule (compound 3). Inhibitors 38 and 44 displayed excellent potency, selectivity (GAPDH/MTS CC(50)), PK parameters in all species studied, and cross genotype activity.