Leucine-rich diet supplementation modulates foetal muscle protein metabolism impaired by Walker-256 tumour

Background

Cancer-cachexia induces a variety of metabolic disorders of protein turnover and is more pronounced when associated with pregnancy. Tumour-bearing pregnant rats have impaired protein balance, which decreases protein synthesis and increases muscle breakdown. Because branched-chain amino acids, especially leucine, stimulate protein synthesis, we investigated the effect of a leucine-rich diet on protein metabolism in the foetal gastrocnemius muscles of tumour-bearing pregnant rats.

Methods

Foetuses of pregnant rats with or without Walker 256 tumours were divided into six groups. During the 20 days of the experiment, the pregnant groups were fed with either a control diet (C, control rats; W, tumour-bearing rats; Cp, rats pair-fed the same normoprotein-diet as the W group) or with a leucine-rich diet (L, leucine rats; LW, leucine tumour-bearing rats; and Lp, rats pair-fed the same leucine-rich diet as the LW group). After the mothers were sacrificed, the foetal gastrocnemius muscle samples were resected, and the protein synthesis and degradation and tissue chymotrypsin-like, cathepsin and calpain enzyme activities were assayed. The muscle oxidative enzymes (catalase, glutathione-S-transferase and superoxide dismutase), alkaline phosphatase enzyme activities and lipid peroxidation (malondialdehyde) were also measured.

Results

Tumour growth led to a reduction in foetal weight associated with decreased serum protein, albumin and glucose levels and low haematocrit in the foetuses of the W group, whereas in the LW foetuses, these changes were less pronounced. Muscle protein synthesis (measured by L-[3H]-phenylalanine incorporation) was reduced in the W foetuses but was restored in the LW group. Protein breakdown (as assessed by tyrosine release) was enhanced in the L and W groups, but chymotrypsin-like activity increased only in group W and tended toward an increase in the LW foetuses. The activity of cathepsin H was significantly higher in the W group foetuses, but the proteolytic calcium-dependent pathway showed similar enzyme activity. In parallel, an intense oxidative stress process was observed only in the group W foetuses.

Conclusions

These data suggested that the proteasomal and lysosomal proteolytic pathways and oxidative stress are likely to participate in the process of foetal muscle catabolism of Walker’s tumour-bearing pregnant rats. The present work shows that foetal muscle can be protected by supplementation with a leucine-rich diet.     

A substrate ambiguous enzyme facilitates genome reduction in an intracellular symbiont

Background

Genome evolution in intracellular microbial symbionts is characterized by gene loss, generating some of the smallest and most gene-poor genomes known. As a result of gene loss these genomes commonly contain metabolic pathways that are fragmented relative to their free-living relatives. The evolutionary retention of fragmented metabolic pathways in the gene-poor genomes of endosymbionts suggests that they are functional. However, it is not always clear how they maintain functionality. To date, the fragmented metabolic pathways of endosymbionts have been shown to maintain functionality through complementation by host genes, complementation by genes of another endosymbiont and complementation by genes in host genomes that have been horizontally acquired from a microbial source that is not the endosymbiont. Here, we demonstrate a fourth mechanism.

Results

We investigate the evolutionary retention of a fragmented pathway for the essential nutrient pantothenate (vitamin B5) in the pea aphid, Acyrthosiphon pisum endosymbiosis with Buchnera aphidicola. Using quantitative analysis of gene expression we present evidence for complementation of the Buchnera pantothenate biosynthesis pathway by host genes. Further, using complementation assays in an Escherichia coli mutant we demonstrate functional replacement of a pantothenate biosynthesis enzyme, 2-dehydropantoate 2-reductase (E.C. 1.1.1.169), by an endosymbiont gene, ilvC, encoding a substrate ambiguous enzyme.

Conclusions

Earlier studies have speculated that missing enzyme steps in fragmented endosymbiont metabolic pathways are completed by adaptable endosymbiont enzymes from other pathways. Here, we experimentally demonstrate completion of a fragmented endosymbiont vitamin biosynthesis pathway by recruitment of a substrate ambiguous enzyme from another pathway. In addition, this work extends host/symbiont metabolic collaboration in the aphid/Buchnera symbiosis from amino acid metabolism to include vitamin biosynthesis.

     

Influence of the selective oestrogen receptor modulator (raloxifene hydrochloride) on IL-6, TNF-alpha, TGF-beta1 and bone turnover markers in the treatment of postmenopausal osteoporosis

BACKGROUND: Osteoporosis that is encountered frequently in postmenopausal women, may cause an increased incidence of vertebral and iliac fractures that are associated with excess morbidity. Raloxifene hydrochloride, a selective oestrogen receptor modulator, has been shown to increase bone mineral density and decrease biochemical markers of bone turnover in postmenopausal women, without stimulatory effects on breast or uterus. Levels of proinflammatory cytokines, including IL-6, and TNF-alpha and TGF-beta1 which are important cytokines involved in remodeling, have been evaluated previously in in vitro studies of osteoporosis. However, there seems to be a paucity of in vivo research concerned with changes in these cytokines in osteoporosis. OBJECTIVE: In this study, we evaluated the effects of raloxifene (Evista); Lilly Pharmaceutical Co. USA, 60 mg/day) on biochemical bone turnover markers, serum parathyroid hormone, and 25-OH vitamin D, as well as the serum levels of IL-6, TNF-alpha and TGF-beta1, in 22 postmenopausal, osteoporotic women before and after 12 weeks of raloxifene treatment. METHODS: Well-matched, postmenopausal, non-osteoporotic control subjects were also enrolled in the study. Serum levels of all the parameters were measured in postmenopausal, osteoporotic women at baseline and end of the study. RESULTS: It was found that serum osteocalcin and parathyroid hormone, and urine deoxypyridinoline levels decreased to normal levels with treatment. Serum 25-OH vitamin D levels after treatment in the patient group were higher than those in the control group. Serum IL-6, TNF-alpha and TGF-beta1 levels did not change significantly with treatment. However, serum levels of IL-6 and TGF-beta1 in the patient group after treatment, decreased to levels lower than those found in the control group. Serum TNF-alpha levels in the patient group before and after treatment, were lower than those in the control group. CONCLUSION: Raloxifene treatment reduces bone turnover biochemical markers, parathyroid hormone and induces 25-OH vitamin D in postmenopausal women. Moreover, it also affects some serum cytokine levels in the postmenopausal period.     

The Deuterator: software for the determination of backbone amide deuterium levels from H/D exchange MS data

Background  

The combination of mass spectrometry and solution phase amide hydrogen/deuterium exchange (H/D exchange) experiments is an effective method for characterizing protein dynamics, and protein-protein or protein-ligand interactions. Despite methodological advancements and improvements in instrumentation and automation, data analysis and display remains a tedious process. The factors that contribute to this bottleneck are the large number of data points produced in a typical experiment, each requiring manual curation and validation, and then calculation of the level of backbone amide exchange. Tools have become available that address some of these issues, but lack sufficient integration, functionality, and accessibility required to address the needs of the H/D exchange community. To date there is no software for the analysis of H/D exchange data that comprehensively addresses these issues.     

Hes1 Increases the Invasion Ability of Colorectal Cancer Cells via the STAT3-MMP14 Pathway

The Notch pathway contributes to self-renewal of tumor-initiating cell and inhibition of normal colonic epithelial cell differentiation. Deregulated expression of Notch1 and Jagged1 is observed in colorectal cancer. Hairy/enhancer of split (HES) family, the most characterized targets of Notch, involved in the development of many cancers. In this study, we explored the role of Hes1 in the tumorigenesis of colorectal cancer. Knocking down Hes1 induced CRC cell senescence and decreased the invasion ability, whereas over-expression of Hes1 increased STAT3 phosphorylation activity and up-regulated MMP14 protein level. We further explored the expression of Hes1 in human colorectal cancer and found high Hes1 mRNA expression is associated with poor prognosis in CRC patients. These findings suggest that Hes1 regulates the invasion ability through the STAT3-MMP14 pathway in CRC cells and high Hes1 expression is a predictor of poor prognosis of CRC.     

Molecular ecology and phylogeography of the bent-wing bat complex (Miniopterus schreibersii) (Chiroptera: Vespertilionidae) in Asia Minor and adjacent regions

In this study we investigate population genetic structure and phylogeography of the bent-wing bat complex ( Miniopterus schreibersii ) in Asia Minor and adjacent regions. PCR amplification and sequencing of the first hypervariable domain of the mitochondrial control region were used to obtain the genetic data. Morphometric differentiation between lineages was analysed by comparing forearm lengths. We found two reciprocally monophyletic lineages within the M . schreibersii complex, identified as M.   s. schreibersii and M. s . pallidus . Distributions of the lineages were allopatric with a U-shaped suture zone passing through Central Anatolia. The suture zone separated coastal regions occupied by M.   s. schreibersii from inland, higher altitude regions occupied by M.   s. pallidus . The lineages showed a considerable sequence divergence of c . 9%, accompanied by a corresponding difference in forearm length. The presence of the genetically distinct lineages, with allopatric distribution and corresponding morphometric differences, probably reflects their long isolation during the ice-age in the Balkans and the Caspian/Caucasus refugia, followed subsequently by expansion into different habitats. Based on the present data, the lineages can be recognized as evolutionary significant units.     

Gem1 and ERMES do not directly affect phosphatidylserine transport from ER to mitochondria or mitochondrial inheritance

In yeast, a protein complex termed the ER-Mitochondria Encounter Structure (ERMES) tethers mitochondria to the endoplasmic reticulum. ERMES proteins are implicated in a variety of cellular functions including phospholipid synthesis, mitochondrial protein import, mitochondrial attachment to actin, polarized mitochondrial movement into daughter cells during division, and maintenance of mitochondrial DNA (mtDNA). The mitochondrial-anchored Gem1 GTPase has been proposed to regulate ERMES functions. Here, we show that ERMES and Gem1 have no direct role in the transport of phosphatidylserine (PS) from the ER to mitochondria during the synthesis of phosphatidylethanolamine (PE), as PS to PE conversion is not affected in ERMES or gem1 mutants. In addition, we report that mitochondrial inheritance defects in ERMES mutants are a secondary consequence of mitochondrial morphology defects, arguing against a primary role for ERMES in mitochondrial association with actin and mitochondrial movement. Finally, we show that ERMES complexes are long-lived, and do not depend on the presence of Gem1. Our findings suggest that the ERMES complex may have primarily a structural role in maintaining mitochondrial morphology.