Sperm Chromatin Immaturity Observed in Short Abstinence Ejaculates Affects DNA Integrity and Longevity In Vitro

BackgroundThe influence of ejaculatory abstinence (EA) on semen parameters and subsequent reproductive outcome is still debatable; hence understanding the impact of EA on sperm structural and functional integrity may provide a valuable information on predicting successful clinical outcome.ObjectiveTo understand the influence of EA on sperm chromatin maturity, integrity, longevity and global methylation status.MethodsThis experimental prospective study included 76 ejaculates from 19 healthy volunteers who provided ejaculates after observing 1, 3, 5 and 7 days of abstinence. Sperm chromatin maturity, DNA integrity and global methylation status were assessed in the neat ejaculate. Sperm motility, DNA integrity and longevity were assessed in the processed fraction of the fresh and frozen-thawed ejaculates to determine their association with the length of EA.ResultsSpermatozoa from 1 day ejaculatory abstinence (EA-1) displayed significantly higher level of sperm chromatin immaturity in comparison to EA-3 (P < 0.05) and EA-5 (P < 0.01) whereas; the number of 5-methyl cytosine immunostained spermatozoa did not vary significantly across groups. On the other hand, in vitro incubation of processed ejaculate from EA-1 resulted in approximately 20 and 40 fold increase in the DNA fragmented spermatozoa at the end of 6 and 24h respectively (P < 0.01–0.001).ConclusionUse of short-term EA for therapeutic fertilization would be a clinically valuable strategy to improve the DNA quality. However, use of such spermatozoa after prolonged incubation in vitro should be avoided as it can carry a substantial risk of transmitting DNA fragmentation to the oocytes.     

Acceleration of ethanolic fermentation by zeolites

Summary Ethanolic fermentation of glucose by Saccharomyces cerevisiae was accelerated in presence of Silicalite only when the yeast was highly flocculent, the zeolite lowering floc size. With the sucrose and molasses, however, fermentation was accelerated even when the yeasts have low flocculence, the zeolite enhancing invertase activity of the yeast cells.     

Response of seedlings of radish (Raphanus sativus) to osmotic shock and external hydrostatic pressure

The effects of an osmotic shock (300 m M mannitol, –0. 67 MPa) or/and increased external hydrostatic pressure on seedlings (42-h-germinated seeds) of radish ( Raphanus sativus L. cv. Tondo Rosso Quarantino) were investigated. The osmotic shock did not inhibit H⁺ extrusion and net K^- uptake, and did not affect the levels of malic acid, reducing sugars, sucrose or amino acids or of the energy charge (i. e. the synthesis of energy-rich phosphate bonds), but inhibited the synthesis of proteins. RNA and DNA, measured as incorporation of labelled precursors. When the osmotic shock was applied together with an increased external hydrostatic pressure of the same magnitude (+0. 67 MPa), the same metabolic parameters and the inhibition of synthesis of RNA and DNA were not substantially affected, while the inhibition of protein synthesis was practically reversed and the energy charge decreased; the recovery of protein synthesis was not due to a change in labelled leucine uptake capability. Increased external hydrostatic pressure alone decreased the energy charge without affecting the other parameters considered.
The possibility that protein synthesis activity is directly controlled by cell turgor pressure (internal hydrostatic pressure) is discussed.     

Deciphering mechanistic implications of antimicrobial and antioxidant potentials of certain new dibutyltin(IV) formulations as possible therapeutic options based on DFT and hybrid materials paradigm

Mechanistic implications of antimicrobial and in vitro antioxidant potentials of a set of newly generated nonbridged mononuclear N,O-orthometallated and carboxylate bridged binuclear nonorthometallated dibutyltin(IV) formulations have been investigated. Some of these formulations were screened for their antibacterial and antifungal activities against Escherichia coli and Candida albicans, respectively whereas in vitro antioxidant potential was examined by Ferric reducing antioxidant power (FRAP) assay. Nonbridged mononuclear N,O-orthometallated dibutyltin(IV) formulations were generated by the reactions of Bu₂SnCl₂ with sodium salts of 2-aminophenol/substituted 2-aminophenol and flexible N-protected amino acids in 1:1:1 molar ratio in refluxing dry THF. Plausible structures of these nonbridged mononuclear N,O-orthometallated dibutyltin(IV) formulations containing flexible N-protected amino acids have been suggested on the basis of spectroscopic and mass studies of some representative formulations. Plausible structures suggested on the basis of spectroscopic studies are corroborated by density functional theory (DFT/B3LYP method) (SPARTAN-20) investigation of a representative dibutyltin(IV) complex and the ligands involved in it. The presence of two different classes of organic ligands in this complex provides an opportunity to study optimized topologies, bonding, distortions, optimized energy, and stability of the complex.     

Methods for Assessment of Biodegradability of Plastic Films in Soil

Traditional and novel techniques were tested and compared for their usefulness in evaluating biodegrad-ability claims made for newly formulated “degradable” plastic film products. Photosensitized polyethylene (PE), starch-PE, extensively plasticized polyvinyl chloride (PVC), and polypropylene (PP) films were incorporated into aerobic soil. Biodegradation was measured for 3 months under generally favorable conditions. Carbon dioxide evolution, residual weight recovery, and loss of tensile strength measurements were supplemented, for some films, by gas chromatographic measurements of plasticizer loss and gel permeation chromatographic (GPC) measurement of polymer molecular size distribution. Six- and 12-week sunlight exposures of photosensitized PE films resulted in extensive photochemical damage that failed to promote subsequent mineralization in soil. An 8% starch-PE film and the plasticized PVC film evolved significant amounts of CO₂ in biodegradation tests and lost residual weight and tensile strength, but GPC measurements demonstrated that all these changes were confined to the additives and the PE and PVC polymers were not degraded. Carbon dioxide evolution was found to be a useful screening tool for plastic film biodegradation, but for films with additives, polymer biodegradation needs to be confirmed by GPC. Photochemical cross-linking of polymer strands reduces solubility and may interfere with GPC measurements of polymer degradation.     

Production of IgM specific recombinant dengue multiepitope protein for early diagnosis of dengue infection

Dengue virus infections have recently undergone dramatic expansion in range, affecting several tropical and subtropical regions of the world. Early detection of dengue infection based on the identification of antibodies has emerged as a practical and reliable means of diagnosis of dengue fever. The recombinant dengue multiepitope (rDME-M) protein specific to IgM in E. coli was produced in a 5-L fermentor for use in diagnostic purpose. After fermentation, dry cell weight was approximately 11.8 g/L of the culture. The rDME-M protein was purified under denaturing conditions using single-step nickel nitrilotriacetate (Ni-NTA) affinity chromatography. The final yield of purified rDME-M protein from this method was approximately 68.5 mg/L of the culture. The purity of rDME-M protein was checked by SDS-PAGE analysis, and the reactivity of this protein was further checked by Western blotting and enzyme-linked immunosorbent assay (ELISA). The purified protein was used as an antigen in the development of an in-house dipstick ELISA and evaluated with a panel of 80 patient sera, characterized using commercially available tests for detection of dengue antibody. The results were in excellent agreement with those of IgM capture ELISA (Pan-Bio) and rapid immunochromatography (IC) test (Pan-Bio). These results show that the in-house dipstick ELISA using rDME-M protein can be used as a promising kit because of its comparable sensitivity, specificity, field applicability, and low cost.     

Neural stem cells as biological minipumps: a faster route to cell therapy for the CNS?

One strategy for the use of neural stem cells (NSCs) in treating neurological disorders is as transplantable "biological minipumps", in which genetically engineered neural stem cells serve as sources of secreted therapeutic (neuroprotective or tumoricidal) agents. Neural stem cells are highly mobile within the brain and demonstrate a tropism for various types of central nervous system (CNS) pathology, making them promising candidates for targeted gene delivery vehicles. Although neural stem cells have also been proposed as a potential source of replacement neurons and astrocytes to repopulate injured or degenerating neural circuits, the challenges involved in rebuilding damaged brain architecture are substantial and remain an active area of investigation. In contrast, the use of NSCs as biological minipumps does not rely on neuronal differentiation, axonal targeting, or synaptogenesis. This strategy may be a faster route to cell-based therapy of the CNS and is poised to move into human clinical trials. This review considers two types of neurologic disease that may be suitable targets for this alternative approach to NSC therapy: glial brain tumors and traumatic brain injury. We examine some of the key scientific and technical issues that must be addressed for the successful use of NSCs as minipumps.     

Telomere dysfunction suppresses spontaneous tumorigenesis in vivo by initiating p53-dependent cellular senescence

Dysfunctional telomeres induce p53-dependent cellular senescence and apoptosis, but it is not known which function is more important for tumour suppression in vivo. We used the p53 ( R172P ) knock-in mouse, which is unable to induce apoptosis but retains intact cell-cycle arrest and cellular senescence pathways, to show that spontaneous tumorigenesis is potently repressed in Terc -/- p53 ( R172P ) mice. Tumour suppression is accompanied by global induction of p53, p21 and the senescence marker senescence-associated-beta-galactosidase. By contrast, cellular senescence was unable to suppress chemically induced skin carcinomas. These results indicate that suppression of spontaneous tumorigenesis by dysfunctional telomeres requires the activation of the p53-dependent cellular senescence pathway.     

Electroporation of cDNA/Morpholinos to targeted areas of embryonic CNS in <Emphasis Type="Italic">Xenopus</Emphasis>

Background  

Blastomere injection of mRNA or antisense oligonucleotides has proven effective in analyzing early gene function in Xenopus. However, functional analysis of genes involved in neuronal differentiation and axon pathfinding by this method is often hampered by earlier function of these genes during development. Therefore, fine spatio-temporal control of over-expression or knock-down approaches is required to specifically address the role of a given gene in these processes.