Angiotensin II type 1 receptor blockade attenuates TGF-beta-induced failure of muscle regeneration in multiple myopathic states

Skeletal muscle has the ability to achieve rapid repair in response to injury or disease. Many individuals with Marfan syndrome (MFS), caused by a deficiency of extracellular fibrillin-1, exhibit myopathy and often are unable to increase muscle mass despite physical exercise. Evidence suggests that selected manifestations of MFS reflect excessive signaling by transforming growth factor (TGF)-beta (refs. 2,3). TGF-beta is a known inhibitor of terminal differentiation of cultured myoblasts; however, the functional contribution of TGF-beta signaling to disease pathogenesis in various inherited myopathic states in vivo remains unknown. Here we show that increased TGF-beta activity leads to failed muscle regeneration in fibrillin-1-deficient mice. Systemic antagonism of TGF-beta through administration of TGF-beta-neutralizing antibody or the angiotensin II type 1 receptor blocker losartan normalizes muscle architecture, repair and function in vivo. Moreover, we show TGF-beta-induced failure of muscle regeneration and a similar therapeutic response in a dystrophin-deficient mouse model of Duchenne muscular dystrophy.     

Isolation murine mesenchymal stem cells by positive selection

Isolation and purification of mesenchymal stem cells (MSCs) from mouse via plastic adherent cultures is arduous because of the unwanted growth of hematopoietic cells and non-MSCs. In this work, homogenous populations of CD34⁺ MSCs from mouse bone marrow were isolated via positive selection. For this purpose, C57Bl/6 mice were killed and bone marrow cells were aspirated before incubation with magnetic bead conjugated to anti-CD34 antibody. A sample of positively selected CD34⁺ cells were prepared for flow cytometry to examine the expression of CD34 antigen and others were subcultured in a 25-cm² culture flask. To investigate the mesenchymal nature, the plastic adherent cultivated cells were induced to differentiate along osteoblastic and adipogenic lineages. Furthermore, the expression of some surface markers was investigated by flow cytometry. According to the result, purified populations of fibroblast-like CD34⁺ cells were achieved in the first passage (1 wk after culture initiation). The cells expressed CD34, CD44, Sca-1, and Vcam-1 antigens (markers) but not CD11b and CD45. They were capable of differentiating into osteocytes and adipocytes. This study indicated that our protocol can result in the efficient isolation of homogenous populations of MSCs from C57BL/6 mouse bone marrow. We have shown that murine bone marrow-derived CD34⁺ cells with plastic adherent properties and capability of differentiating into skeletal lineages in vitro are MSCs.     

Enhanced cadmium-induced testicular necrosis and renal proximal tubule damage caused by gene-dose increase in a Slc39a8-transgenic mouse line

Resistance to cadmium (Cd)-induced testicular necrosis is an autosomal recessive trait defined as the Cdm locus. Using positional cloning, we previously identified the Slc39a8 (encoding an apical-surface ZIP8 transporter protein) as the gene most likely responsible for the phenotype. In situ hybridization revealed that endothelial cells of the testis vasculature express high ZIP8 levels in two sensitive inbred mouse strains and negligible amounts in two resistant strains. In the present study, we isolated a 168.7-kb bacterial artificial chromosome (BAC), carrying only the Slc39a8 gene, from a Cd-sensitive 129/SvJ BAC library and generated BAC-transgenic mice. The BTZIP8-3 line, having three copies of the 129/SvJ Slc39a8 gene inserted into the Cd-resistant C57BL/6J genome (having its normal two copies of the Slc39a8 gene), showed tissue-specific ZIP8 mRNA expression similar to wild-type mice, mainly in lung, testis, and kidney. The 2.5-fold greater expression paralleled the fact that the BTZIP8-3 line has five copies, whereas wild-type mice have two copies, of the Slc39a8 gene. The ZIP8 mRNA and protein localized especially to endothelial cells of the testis vasculature in BTZIP8-3 mice. Cd treatment reversed Cd resistance (seen in nontransgenic littermates) to Cd sensitivity in BTZIP8-3 mice; reversal of the testicular necrosis phenotype confirms that Slc39a8 is unequivocally the Cdm locus. ZIP8 also localized specifically to the apical surface of proximal tubule cells in the BTZIP8-3 kidney. Cd treatment caused acute renal failure and signs of proximal tubular damage in the BTZIP8-3 but not nontransgenic littermates. BTZIP8-3 mice should be a useful model for studying Cd-induced disease in kidney. kidney; testis; ZIP8; bacterial artificial chromosome     

Spermidine/spermine N1-acetyltransferase overexpression in kidney epithelial cells disrupts polyamine homeostasis, leads to DNA damage, and causes G2 arrest

Expression of spermidine/spermine N¹-acetyltransferase (SSAT) increases in kidneys subjected to ischemia-reperfusion injury (IRI). Increased expression of SSAT in vitro leads to alterations in cellular polyamine content, depletion of cofactors and precursors of polyamine synthesis, and reduced cell proliferation. In our model system, a >28-fold increase in SSAT levels in HEK-293 cells leads to depletion of polyamines and elevation in the enzymatic activities of ornithine decarboxylase and S-adenosylmethionine decarboxylase, suggestive of a compensatory reaction to increased polyamine catabolism. Increased expression of SSAT also led to DNA damage and G₂ arrest. The increased DNA damage was primarily due to the depletion of polyamines. Other factors such as increased production of H₂O₂ due to polyamine oxidase activity may play a secondary role in the induction of DNA lesions. In response to DNA damage the ATM/ATR Chk1/2 DNA repair and cell cycle checkpoint pathways were activated, mediating the G₂ arrest in SSAT-expressing cells. In addition, the activation of ERK1 and ERK2, which play integral roles in the G₂/M transition, is impaired in cells expressing SSAT. These results indicate that the disruption of polyamine homeostasis due to enhanced SSAT activity leads to DNA damage and reduced cell proliferation via activation of DNA repair and cell cycle checkpoint and disruption of Raf MEK ERK pathways. We propose that in kidneys subjected to IRI, one mechanism through which increased expression of SSAT may cause cellular injury and organ damage is through induction of DNA damage and the disruption of cell cycle. ischemia-reperfusion injury; polyamine depletion; cell proliferation; DNA repair; cell cycle arrest     

Biodesulfurization of refractory organic sulfur compounds in fossil fuels

The stringent new regulations to lower sulfur content in fossil fuels require new economic and efficient methods for desulfurization of recalcitrant organic sulfur. Hydrodesulfurization of such compounds is very costly and requires high operating temperature and pressure. Biodesulfurization is a non-invasive approach that can specifically remove sulfur from refractory hydrocarbons under mild conditions and it can be potentially used in industrial desulfurization. Intensive research has been conducted in microbiology and molecular biology of the competent strains to increase their desulfurization activity; however, even the highest activity obtained is still insufficient to fulfill the industrial requirements. To improve the biodesulfurization efficiency, more work is needed in areas such as increasing specific desulfurization activity, hydrocarbon phase tolerance, sulfur removal at higher temperature, and isolating new strains for desulfurizing a broader range of sulfur compounds. This article comprehensively reviews and discusses key issues, advances and challenges for a competitive biodesulfurization process.     

Effect of feeding and sludge age on acclimated bacterial community and fate of slowly biodegradable substrate

The study investigated the effect of feeding regime and sludge age on starch utilization. For this purpose, parallel sequencing batch reactors were operated with pulse and continuous feeding of soluble starch at sludge ages of 8 and 2 days. Pulse feeding induced almost complete conversion of starch to glycogen, while storage was lowered and accompanied with direct growth under continuous feeding, regardless of sludge age. Low sludge age did not alter simultaneous storage and utilization for direct growth but it slightly favoured direct utilization due to faster growing biomass. Experimental results suggested adsorption of starch onto biomass as a preliminary removal mechanism prior to hydrolysis at sludge age of 8 days. Adsorption was not noticeable as substrate removal, glycogen generation and dissolved oxygen decrease were synchronous at sludge age of 2 days. Bacterial community always included fractions storing glycogen although sludge age only affected the relative magnitude of filamentous growth.     

Immunostimulant effects of Cotinus coggyria on rainbow trout (Oncorhynchus mykiss)

In this study, non-specific immune effects of tetra (Cotinus coggyria) on rainbow trout (Oncorhynchus mykiss) by dietary intake were investigated. Fish were fed daily ad libitum with diets containing 0.5% and 1.0% tetra for 3 weeks. After this period, fish were switched back to the basal diet for additional 6 weeks. Extracellular and intracellular respiratory burst activities, phagocytosis in blood leukocytes, lysozyme activities, and total plasma protein levels were evaluated at the end of the tetra feeding period and every 3 weeks during the basal diet period. Extracellular and intracellular respiratory burst activities, phagocytic activity, lysozyme activity and total protein level parameters of the groups containing 0.5% and 1.0% tetra were higher than the control group at the end of the 3rd, 6th and 9th weeks, respectively (P < 0.05). The highest values of the non-specific immune parameters were observed in the group fed with 1.0% tetra. Tetra groups did not show any significant difference (P > 0.05) in terms of specific growth rate and average weight of the fish.