Methane fermentation of brewery by-products

A laboratory-scale research program was undertaken to investigate the kinetics of the mesophilic (37°C) anaerobic digestion of brewery industry by-product. The purpose was to develop data for the design and operation of full-scale units which could be used to generate methane fuel gas from these materials. This is important because the brewery industry has been susceptible to shortages of natural gas in recent years. The minimum SRT is 2.3 days, although for design purposes as much as ten days is recommended. The biomass yield is 0.512 g volatile suspended solids (VSS)/g volatile solids (VS) or 0.421 g VSS/g chemical oxygen demand (COD). The maintenance requirement is 0.052 g VS/g VSS per day or 0.061 g COD/g VSS per day. The specific methane yield is 2.51 liter/g VSS, and the methane productivity is 0.32–0.41 liter/g dry substrate added or 0.69–0.91 liter/g destroyed. The maximum loading rate for which substrate inhibition is not observed is 6 g dry substrate added per liter per day. The results of the entire program indicate that processing brewery by-product in this manner is both technically feasible and economically attractive.     

Characterization of extremely halophilic Archaea isolated from the Ayvalik Saltern,Turkey

Seven extremely halophilic strains were isolated from the Ayvalik Saltern in the north-eastern part of Turkey. Chemical analyses of the brine and salt samples were performed to measure their salt content, hardness and pH. Isolated strains were tested for their antibiotic sensitivities; cell and colony morphologies; hydrolysis of casein, starch, gelatin, Tween 20 and Tween 80; and oxidase and catalase activity. All strains were found to belong to the domain Archaea. Characterization of polar lipids by thin layer chromatography indicated that all isolates contained phytanyl diether derivatives of phosphatidylglycerol (PG), the methyl ester of phosphatidyl glycerophosphate (PGP-Me), and phosphatidylglycerosulphate (PGS). Four isolates had triglycosyl diether (TGD-2) as glycolipid, and the other three contained a sulphated diglycosyl diether instead. All isolates were examined for the presence of plasmids by agarose gel electrophoresis. Four strains were found to harbour plasmids ranging in size from 13.8 to 15.3 kbp. Correlation between the protein profiles in SDS–PAGE and the phenotypic properties of the strains was poor. The data presented here provide the first published account of the microbiota of the Ayvalik saltern, which provides a large part of the salt produced in Turkey.     

In utero transplantation of neural stem cells ameliorates maternal inflammation-induced prenatal white matter injury

Prenatal white matter injury is a serious problem due to maternal inflammation leading to postnatal disabilities. In this study, we used the periventricular leukomalacia (PVL) model as a common prenatal white matter injury by maternal administration of lipopolysaccharide (LPS). Neural stem cells (NSCs) have shown therapeutic ability in neurological disorders through a different mechanism such as immunomodulation. Here, we studied the preventive potential of NSCs following in utero transplantation into the embryonic lateral ventricle in an LPS-induced white matter injury model. Pregnant animals were divided into three groups and received phosphate buffered saline, LPS, or LPS + NSCs. The brains of offspring were obtained and evaluated by real-time polymerase chain reaction (PCR), immunohistochemy, enzyme-linked immunosorbent assay (ELISA), terminal deoxynucleotidyl transferase-mediated biotinylated-dUTP nick-end labeling (TUNEL), and caspase-3 activity assay. The LPS-induced maternal inflammation degenerated the myelin sheath in the offspring periventricular region which was associated with an increased microglial number, oligodendrocytes degeneration, proinflammatory cytokine secretion, and cell apoptosis. The transplanted NSCs homed into the brain and ameliorated the evaluated parameters. The expression of proinflammatory cytokines interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α), cell apoptosis and caspase-3 activity were inhibited by NSCs. In addition, Olig2 and myelin basic protein immunohistochemy staining showed that prenatal NSCs transplantation augmented the myelination in the periventricular white matter of offspring. In conclusion, we think that prenatal therapeutic strategies, such as in utero NSCs transplantation, may prevent prenatal white matter injury after birth.     

Roles for Kisspeptin in proliferation and differentiation of spermatogonial cells isolated from mice offspring when the cells are cocultured with somatic cells

Kisspeptin (Kp) expression in testis has caused most of the recent research surveying its functional role in this organ. This peptide influences spermatogenesis and sperm capacitation, so it is considered as a regulator of reproduction. Kp roles exert through hypothalamic/pituitary/gonadal axis. We aimed to evaluate direct roles for Kp on proliferation and differentiation of spermatogonial cells (SCs) when the cells are cocultured with somatic cells. Somatic cells and SCs were isolated from adult azoospermic and newborn mice and then enriched using a differential attachment technique. After the evaluation of identity and colonization for SCs, the cells were cocultured with somatic cells, and three doses of Kp (10⁻⁸-10⁻⁶ M) was assessed on proliferation (through evaluation of MVH and ID4 markers) and differentiation (via evaluation of c-Kit and SCP₃, TP_1, TP₂, and, Prm₁ markers) of the coculture system. Investigations were continued for four succeeding weeks. At the end of each level of testosterone in the culture media was also evaluated. We found positive influence from Kp on proliferative and differentiative markers in SCs cocultured with somatic cells. These effects were dose-dependent. There was no effect for Kp on testosterone level. From our findings, we simply conclude that Kp as a neuropeptide for influencing central part of reproductive axis could also positively affect peripheral processes related to spermatogenesis without having an effect on steroidogenesis.     

A novel VHH nanobody against the active site (the CA domain) of tumor-associated, carbonic anhydrase isoform IX and its usefulness for cancer diagnosis

Expression of carbonic anhydrase IX (CAIX) significantly increases under hypoxic conditions in tumor cells. CAIX activity is executed by the catalytic domain (CA) located on the extracellular part of the enzyme. Neutralization of CAIX enzymatic activity reduces malignancy and survival of tumor cells. To inhibit the enzymatic activity, a VHH nanobody was developed against the CA domain of CAIX using phage display technology. Following immunization of a camel with the recombinant CAIX, VHH fragments were isolated by nested PCR on lymphocyte cDNA. Binding affinity of isolated nanobodies was tested by ELISA. A clone (K24) with the highest binding affinity was expressed in a soluble form. Affinity of K24 nanobody was determined to be approx. 2.3 × 10^?5. K24 nanobody recognized the expressed CAIX in the HeLa cell lines with high selectivity and specificity. These findings thus have usefulness for the diagnosis and treatment of cancers.     

Functional characterization of two RAB27A missense mutations found in Griscelli syndrome type 2

Human Griscelli syndrome type 2 (GS-2) is characterized by partial albinism and a severe immunologic disorder as a result of RAB27A mutations. In melanocytes, Rab27A forms a tripartite complex with a specific effector Slac2-a/melanophilin and myosin Va, and the complex regulates melanosome transport. Here, we report a novel homozygous missense mutation of Rab27A, i.e. K22R, in a Persian GS-2 patient and the results of analysis of the impact of the K22R mutation and the previously reported I44T mutation on protein function. Both mutations completely abolish Slac2-a/melanophilin binding activity but they affect the biochemical properties of Rab27A differently. The Rab27A(K22R) mutant lacks the GTP binding ability and exhibits cytosolic localization in melanocytes. By contrast, neither intrinsic GTPase activity nor melanosomal localization of Rab27A is affected by the I44T mutation, but the Rab27A(I44T) mutant is unable to recruit Slac2-a/melanophilin. Interestingly, the two mutations differently affect binding to other Rab27A effectors, Slp2-a, Slp4-a/granuphilin-a, and Munc13-4. The Rab27A(K22R) mutant normally binds Munc13-4, but not Slp2-a or Slp4-a, whereas the Rab27A(I44T) mutant shows reduced binding activity to Slp2-a and Munc13-4 but normally binds Slp4-a.     

PPAR-γ: Its ligand and its regulation by microRNAs

Peroxisome proliferator-activated receptors (PPARs) belong to the nuclear receptor superfamily. PPARs are categorized into three subtypes, PPARα, β/δ, and γ, encoded by different genes, expressed in diverse tissues and participate in various biological functions and can be activated by their metabolic derivatives in the body or dietary fatty acids. The PPAR-γ also takes parts in the regulation of energy balance, lipoprotein metabolism, insulin sensitivity, oxidative stress, and inflammatory signaling. It has been implicated in the pathology of numerous diseases including obesity, diabetes, atherosclerosis, and cancers. Among various cellular and molecular targets that are able to regulate PPAR-γ and its underlying pathways, microRNAs (miRNAs) appeared as important regulators. Given that the deregulation of these molecules via targeting PPAR-γ could affect initiation and progression of various diseases, identification of miRNAs that affects PPAR-γ could contribute to the better understanding of roles of PPAR-γ in various biological and pathological conditions. Here, we have summarized the function and various ligands of PPAR-γ and have highlighted various miRNAs involved in the regulation of PPAR-γ.