Dysferlin and muscle membrane repair

The ability to repair membrane damage is conserved across eukaryotic cells and is necessary for the cells to survive a variety of physiological and pathological membrane disruptions. Membrane repair is mediated by rapid Ca(2+)-triggered exocytosis of various intracellular vesicles, such as lysosomes and enlargeosomes, which lead to the formation of a membrane patch that reseals the membrane lesion. Recent findings suggest a crucial role for dysferlin in this repair process in muscle, possibly as a Ca(2+) sensor that triggers vesicle fusion. The importance of membrane repair is highlighted by the genetic disease, dysferlinopathy, in which the primary defect is the loss of Ca(2+)-regulated membrane repair due to dysferlin deficiency. Future research on dysferlin and its interacting partners will enhance the understanding of this important process and provide novel avenues to potential therapies.     

Dysferlin forms a dimer mediated by the C2 domains and the transmembrane domain in vitro and in living cells

Dysferlin was previously identified as a key player in muscle membrane repair and its deficiency leads to the development of muscular dystrophy and cardiomyopathy. However, little is known about the oligomerization of this protein in the plasma membrane. Here we report for the first time that dysferlin forms a dimer in vitro and in living adult skeletal muscle fibers isolated from mice. Endogenous dysferlin from rabbit skeletal muscle exists primarily as a ～460 kDa species in detergent-solubilized muscle homogenate, as shown by sucrose gradient fractionation, gel filtration and cross-linking assays. Fluorescent protein (YFP) labeled human dysferlin forms a dimer in vitro, as demonstrated by fluorescence correlation spectroscopy (FCS) and photon counting histogram (PCH) analyses. Dysferlin also dimerizes in living cells, as probed by fluorescence resonance energy transfer (FRET). Domain mapping FRET experiments showed that dysferlin dimerization is mediated by its transmembrane domain and by multiple C2 domains. However, C2A did not significantly contribute to dimerization; notably, this is the only C2 domain in dysferlin known to engage in a Ca-dependent interaction with cell membranes. Taken together, the data suggest that Ca-insensitive C2 domains mediate high affinity self-association of dysferlin in a parallel homodimer, leaving the Ca-sensitive C2A domain free to interact with membranes.     

Effects of bone marrow mesenchymal stromal cells on gross motor function measure scores of children with cerebral palsy: a preliminary clinical study

Background aimsPre-clinical evidence indicates that autologous bone marrow-derived mesenchymal stromal cell (BM-MSC) transplantation improves motor function in patients with central nervous system disorders.MethodsAfter providing informed consent, 52 patients with cerebral palsy (CP) who met the study criteria received BM-MSC transplantation. Gross motor function was assessed using the Gross Motor Function Measure (GMFM)-88 and GMFM-66 scales at baseline (before transplantation) and at 1 month, 6 months and 18 months post-transplantation. The participants completed the trial without visible side effects. The GMFM-66 percentile (motor growth curves) was used as the control index of motor function to exclude the interference of improvement with age.ResultsThe score domains A, B, C and D and the total GMFM-88 and GMFM-66 scores in participants increased at 1 month, 6 months and 18 months post-transplantation compared with the baseline value (P < 0.01). The scores of domain E also increased at 6 months and 18 months post-transplantation, although they were not significantly increased at 1 month post-transplantation. There were significant increases in the GMFM-66 score and the GMFM-66 percentile corresponding to patient age and Gross Motor Function Classification System level after cell transplantation.ConclusionsAutologous BM-MSC transplantation appears to be a feasible, safe and effective therapy for patients with CP. The treatment improved the development of children with CP with regard to motor function.     

类产碱假单胞菌谷氨酸脱氢酶的提纯、鉴定及某些特性的初步研究

从类产碱假单胞菌纯化出电泳纯的谷氨酸脱氢酶,用聚丙烯酰胺梯度凝胶电泳和ＳＤＳ－聚丙烯酰胺凝胶电泳测得分子量为２９０ ｋＤ,亚基分子量为４７ ｋＤ,提示该酶为六聚体．该酶对ＮＡＤＰ（Ｈ）和底物均具有高度专一性,对谷氨酸、α－酮戊二酸及ＮＡＤＰ＋ 的Ｋｍ 值分别为：２８ ｍ ｍ ｏｌ／Ｌ、１．２ｍ ｍ ｏｌ／Ｌ及０．０６３ ｍ ｍ ｏｌ／Ｌ．用Ｈｉｌｌ作图法求得酶对ＮＨ＋４ 和ＮＡＤＰＨ 的［Ｓ］０．５分别为２４ ｍ ｍ ｏｌ／Ｌ和０．０３７ ｍ ｍ ｏｌ／Ｌ．最适反应温度为５０℃,催化氨化反应和脱氨反应的最适ｐＨ 分别为８．０和８．８,在热稳定性方面不及嗜热细菌的谷氨酸脱氢酶稳定．提纯的谷氨酸脱氢酶在低温（４℃）条件下,可在Ｔｒｉｓ－ＨＣｌ缓冲液中贮存半年以上,活力无明显下降,冷冻则可导致纯酶液迅速失活．氮源对菌体谷氨酸脱氢酶水平有显著影响．     

保护性耕作条件下旱地农田麦-豆双序列轮作体系的水分动态及产量效应

保护性耕作(conservationtillage)能够减少水土流失、提高耕地产量,是一类具有生态保护意义的持续性农业耕作形式。2002年至2004年在定西旱地农业地区进行了保护性耕作条件下旱地农田春小麦豌豆双序列轮作土壤水分动态及产量效应的试验研究,结果表明:保护性耕作能够显著改善0～200cm土层土壤贮水量及含水量,随着降水量的增多土壤对降水的保蓄能力增强。在降水较少年份免耕秸秆覆盖的这种作用表现突出,而在降水充沛的年份免耕地膜覆盖则更具优势。耕层土壤水分因受降水等因素的影响而变化剧烈,耕层以下土壤水分变幅相对较小。播种期、五叶期及收获期土壤具有较高含水量,而开花期土壤含水量则较低。在两种轮作体系中,播种期春小麦和豌豆免耕秸秆覆盖处理0～50cm土层含水量分别较常规耕作增加28%、26%和11%、23%,降水生产效率较常规耕作提高了17.79%～26.81%。在春小麦豌豆轮作体系中免耕秸秆覆盖处理的作物产量(春小麦 豌豆)及水分利用效率分别为3420kghm2和8.11kg(hm2·mm),较常规耕作分别提高26.81%和25.39%。     

Land eco-security evaluation in the Shule River Basin during 2005–2014 in Gansu Province,China

The Shule River Basin is an ecologically fragile area in arid zone. To understand the state of land eco-security, the Environment-Economic-Society model was applied to build a land eco-security evaluation index system in the Shule River Basin. An entropy-weighted and matter-element model was built for eco-security evaluation from 2005 to 2014. Principal component analysis was used to quantitatively study the limiting factors of land ecological security. The result showed: the direction of development of the state of land eco-security in the Shule River Basin from 2005 to 2014 was characterized by “unsafe (No₄) → safe (No₁),” and presented an upward trend. The land eco-security status during 2005–2007 was in the “Unsafe” state and the state changed to “Critical Safe” in 2008–2009, “Safer” in 2010–2011, but “Safe” in 2012–2014. The key factors that affected land eco-security in the Shule River Basin were Per Capita Arable Land, Forest Cover Rate, Per Capita Water Resources, Water Production Modulus, the Tertiary Industry Output Value, and GDP Ratio and Water Consumption. Among them, Forest Cover Rate and Water Production Modulus had the greatest impact, with principal component loads of up to 0.973 and 0.968, respectively. The result of this study is expected to serve as reference and support for the conservation and management of Shule River Basin to ensure sustainable development.     

RNA interference-mediated silencing of iASPP induces cell proliferation inhibition and G0/G1 cell cycle arrest in U251 human glioblastoma cells

iASPP is an evolutionally conserved inhibitory member of the ASPP (apoptosis-stimulating protein of p53) protein family. Overexpression of iASPP was observed in several types of human tumors, however, its role in tumorigenesis has not been fully clarified. To investigate the role of iASPP in human glioblastoma multiforme (GMB) progression, the authors employed lentivirus-mediated shRNA to silence endogenous iASPP expression and elucidated iASPP function by analysis of viability, colony formation, DNA synthesis, and cell cycle in p53-mutant glioblastoma cell line U251. iASPP was significantly and sustainably knocked down by iASPP-specific shRNA in U251 cells. Stable down-regulation of iASPP expression-induced cell proliferation inhibition and G0/G1 cell cycle arrest by down-regulation of cyclin D1 and up-regulation of p21(Waf1/Cip1). Thus, the findings not only provide a molecular basis for the role of iASPP in cell cycle progression of glioblastoma cells but also suggest a novel therapeutic target for the treatment of GBM.     

Measurement of sub-membrane [Ca2+] in adult myofibers and cytosolic [Ca2+] in myotubes from normal and mdx mice using the Ca2+ indicator FFP-18

The hypothesis that intracellular Ca(2+) is elevated in dystrophic (mdx) skeletal muscle due to increased Ca(2+) influx is controversial. As the sub-sarcolemmal Ca(2+) ([Ca(2+)](mem)) should be even higher than the global cytosolic Ca(2+) in the presence of increased Ca(2+) influx, we investigated [Ca(2+)](mem) levels in collagenase-isolated adult flexor digitorum brevis (FDB) myofibres and myotubes of mdx and normal mice with the near-membrane Ca(2+) indicator FFP-18. Confocal imaging showed strong localization of FFP-18 to the sarcolemma only. No significant difference in [Ca(2+)](mem) was found in FDB myofibres of normal (77.3+/-3.8 nM, n=68) and mdx (79.3+/-5.6 nM, n=21, p=0.89) mice using FFP-18. Increasing external Ca(2+) to 18 mM did not significantly affect [Ca(2+)](mem) in either the normal or mdx myofibres. In the myotubes, the FFP-18 was non-selectively incorporated, distributing throughout the cytoplasm, and FFP-18-derived [Ca(2+)] values were similar to values obtained with Fura-2. Nevertheless, in the mdx myotubes, the [Ca(2+)] measured with FFP-18 increased linearly to a level approximately 2.75 times that of controls as the time of culture was prolonged. In older mdx myotubes (>or=8 days in culture), 18 mM extracellular Ca(2+) increased the steady state cytosolic [Ca(2+)] to approximately 22 times greater level than controls. This study suggests that the sub-sarcolemmal Ca(2+) homeostasis is well maintained in isolated adult mdx myofibers and also further supports the hypothesis that cytosolic Ca(2+) handling is compromised in mdx myotubes.     

Some aspects of the growth of Gracilaria tenuistipitata in pond culture

The effect of temperature, salinity, nitrogen, culture density and depth on the growth of Gracilaria tenuistipitata were investigated between April 1985 and March 1986 in outdoor ponds in Guangxi Province, South China. The mean annual growth rate was 2.4% per day. Under favourable temperatures of 20–30 °C, daily growth rate may reach as high as 3.3%. Salinity had an obvious effect on growth and photosynthesis and growth peaked at 21, with a broad plateau between 7–27. Growth experiments showed that a total nitrogen (NH₄-N plus NO₃-N) concentration of 4 M was sufficient to enable the plants to maintain a daily growth rate of 2.7%. The best growth of the plant was obtained at a culture density of 0.5–1 kg m^–2 and a culture depth of 30 cm in the pond.