TLR4 Activation Promotes Bone Marrow MSC Proliferation and Osteogenic Differentiation via Wnt3a and Wnt5a Signaling

Mesenchymal stem cells (MSCs) from adult bone marrow maintain their self-renewal ability and the ability to differentiate into osteoblast. Thus, adult bone marrow MSCs play a key role in the regeneration of bone tissue. Previous studies indicated that TLR4 is expressed in MSCs and is critical in regulating the fate decision of MSCs. However, the exact functional role and underlying mechanisms of how TLR4 regulate bone marrow MSC proliferation and differentiation are unclear. Here, we found that activated TLR4 by its ligand LPS promoted the proliferation and osteogenic differentiation of MSCs in vitro. TLR4 activation by LPS also increased cytokine IL-6 and IL-1β production in MSCs. In addition, LPS treatment has no effect on inducing cell death of MSCs. Deletion of TLR4 expression in MSCs completely eliminated the effects of LPS on MSC proliferation, osteogenic differentiation and cytokine production. We also found that the mRNA and protein expression of Wnt3a and Wnt5a, two important factors in regulating MSC fate decision, was upregulated in a TLR4-dependent manner. Silencing Wnt3a with specific siRNA remarkably inhibited TLR4-induced MSC proliferation, while Wnt5a specific siRNA treatment significantly antagonized TLR4-induced MSC osteogenic differentiation. These results together suggested that TLR4 regulates bone marrow MSC proliferation and osteogenic differentiation through Wnt3a and Wnt5a signaling. These finding provide new data to understand the role and the molecular mechanisms of TLR4 in regulating bone marrow MSC functions. These data also provide new insight in developing new therapy in bone regeneration using MSCs by modulating TLR4 and Wnt signaling activity.     

Genomic constitution and taxonomy of the Chinese hexaploids Elymus cylindricus and E. breviaristatus (Poaceae: Triticeae)

Elymus cylindricus (2n = 6x = 42) and E. breviaristatus (2n = 6x = 42) are distributed in grasslands and deserts of northern and north‐western China. Their genomic constitution and taxonomic status are unclear. Elymus cylindricus was crossed with E. wawawaiensis J.R.Carlson & Barkworth ( StH ), Roegneria grandis Keng ( StY ) and Campeiostachys dahurica (Turcz. ex Griseb.) B.R.Baum, J.L. Y ang & C. Y en var. dahurica ( StYH ). Meiotic pairing in the hybrids E. cylindricus × E. wawawaiensis ( StH ), E. cylindricus × R. grandis ( StY ) and E. cylindricus × C. dahurica var. dahurica ( StYH ) showed on average 10.00, 11.30 and 20.92 bivalents per cell, respectively. Elymus breviaristatus was crossed with C. dahurica var. dahurica ( StYH ) and E. cylindricus. Chromosome pairing in the hybrids of E. breviaristatus × C. dahurica var. dahurica and E. breviaristatus × E. cylindricus showed on average 19.60 and 19.27 bivalents, respectively. Genomic in situ hybridization (GI SH ) revealed the presence of St , Y and H genomes in E. cylindricus and E. breviaristatus. An intergenomic rearrangement was observed in E. cylindricus using GI SH . Meiotic pairing data and GI SH indicated that both E. cylindricus and E. breviaristatus are allohexaploids containing the StYH genomes. Elymus cylindricus and E. breviaristatus should be treated as Campeiostachys dahurica var. cylindrica and Campeiostachys breviaristata, respectively.     

河北张家口康巴诺尔湖国家湿地公园遗鸥繁殖群新发现

2014年4月14日在张家口市康保县康巴诺尔湖国家湿地公园(东经114°35′06″~114°36′32″,北纬41°49′05″~41°50′33″)发现遗鸥(Larus relictus)3 000余只。同年7月在湖边发现新繁殖的遗鸥幼鸟觅食。2015年4月底发现遗鸥在湖心岛筑巢产卵,同年6月采用国产大疆精灵3四轴航拍飞行器航拍法及样方法统计湖心岛遗鸥种群数量,共记录遗鸥成鸟2 100余只,806巢,出壳雏鸟2 080只。此次发现属于我国遗鸥繁殖地的新发现。     

Functional TRPV2 and TRPV4 channels in human cardiac c‐kit+ progenitor cells

The cellular physiology and biology of human cardiac c‐kit⁺ progenitor cells has not been extensively characterized and remains an area of active research. This study investigates the functional expression of transient receptor potential vanilloid (TRPV) and possible roles for this ion channel in regulating proliferation and migration of human cardiac c‐kit⁺ progenitor cells. We found that genes coding for TRPV2 and TRPV4 channels and their proteins are significantly expressed in human c‐kit⁺ cardiac stem cells. Probenecid, an activator of TRPV2, induced an increase in intracellular Ca²⁺ (Ca²⁺_i), an effect that may be attenuated or abolished by the TRPV2 blocker ruthenium red. The TRPV4 channel activator 4α‐phorbol 12‐13‐dicaprinate induced Ca²⁺_i oscillations, which can be inhibited by the TRPV4 blocker RN‐1734. The alteration of Ca²⁺_i by probenecid or 4α‐phorbol 12‐13‐dicprinate was dramatically inhibited in cells infected with TRPV2 short hairpin RNA (shRNA) or TRPV4 shRNA. Silencing TRPV2, but not TRPV4, significantly reduced cell proliferation by arresting cells at the G0/G1 boundary of the cell cycle. Cell migration was reduced by silencing TRPV2 or TRPV4. Western blot revealed that silencing TRPV2 decreased expression of cyclin D1, cyclin E, pERK1/2 and pAkt, whereas silencing TRPV4 only reduced pAkt expression. Our results demonstrate for the first time that functional TRPV2 and TRPV4 channels are abundantly expressed in human cardiac c‐kit⁺ progenitor cells. TRPV2 channels, but not TRPV4 channels, participate in regulating cell cycle progression; moreover, both TRPV2 and TRPV4 are involved in migration of human cardiac c‐kit⁺ progenitor cells.     

EXPRESSION AND EFFECTS OF MUTANT Bombyx mori ACETYLCHOLINESTRASE1 IN BmN CELLS

The main mechanism of toxicity of organophosphate (OP) and carbamate (CB) insecticides is their irreversible binding and inhibition of acetylcholinestrase (AChE), encoded by ace1 (acetylcholinestrase gene 1), leading to eventual death of insects. Mutations in AChE may significantly reduce insects susceptibility to these pesticides. Bombyx mori is an important beneficial insect, and no OP‐ or CB‐resistant strains have been generated. In this study, wild‐type ace1 (wace1) and mutant ace1 (mace1) were introduced into BmN cells, confirmed by screening and identification. The expression of wace1 and mace1 in the cells was confirmed by Western blot and their expression levels were about 21‐fold higher than the endogenous ace1 level. The activities of AChE in wace1 and mace1 transgenic cells were 10.6 and 20.2% higher compared to control cells, respectively. mace1 transgenic cells had higher remaining activity than wace1 transgenic cells under the treatment of physostigmine (a reversible cholinesterase inhibitor) and phoxim (an OP acaricide). The results showed that ace1 transgene can significantly improve ace1 expression, and ace1 mutation at a specific site can reduce the sensitivity to AChE inhibitors. Our study provides a new direction for the exploration of the relationship between AChE mutations and drug resistance.     

A large‐scale phylogeny of the lycophyte genus Selaginella (Selaginellaceae: Lycopodiopsida) based on plastid and nuclear loci

The lycophyte genus Selaginella alone constitutes the family Selaginellaceae, the largest of the lycophyte families. The genus is estimated to contain 700–800 species distributed on all continents except Antarctica, with highest species diversity in tropical and subtropical regions. The monophyly of Selaginella in this broad sense has rarely been doubted, whereas its intrageneric classification has been notoriously contentious. Previous molecular studies were based on very sparse sampling of Selaginella (up to 62 species) and often used DNA sequence data from one genome. In the present study, DNA sequences of one plastid (rbcL) and one nuclear (ITS) locus from 394 accessions representing approximately 200 species of Selaginella worldwide were used to infer a phylogeny using maximum likelihood, Bayesian inference and maximum parsimony methods. The study identifies strongly supported major clades and well resolves relationships among them. Major results include: (i) six deep‐level clades are discovered representing the deep splits of Selaginella; and (ii) 20 major clades representing 20 major evolutionary lineages are identified, which differ from one another in molecular, macro‐morphological, ecological and spore features, and/or geographical distribution.     

Efficient Detection of Leptosphaeria maculans from Infected Seed lots of Oilseed Rape

An efficient DNA extraction protocol and polymerase chain reaction (PCR) assay for detecting Leptosphaeria maculans from infected seed lots of oilseed rape were developed. L. maculans, the causal agent of blackleg, a damaging disease in oilseeds rape/canola worldwide, was listed as a quarantine disease by China in 2009. China imports several millions of tons of oilseeds every year. So there is a high risk that this pathogen will be introduced to China via contaminated seeds. Seed contamination is one of the most significant factors in the global spread of phytopathogens. Detection of L. maculans in infected seed lots by PCR assay is difficult due to the low level of pathogen mycelium/spores on seeds and PCR inhibitors associated with the seeds of oilseed rape. In our study, these two major obstacles were overcome by the development of a two‐step extraction protocol combined with a nested PCR. This extraction protocol (kit extraction after CTAB method) can efficiently extract high‐quality DNA for PCR. Amplification results showed that the detection threshold for conventional PCR and nested PCR was, respectively, 1 ng and 10 fg of DNA per μl in mycelia samples. On contaminated seed lots of oilseed rape, the detection threshold of conventional and nested PCR was 709 fg/μl and 709 ag/μl of DNA, respectively. The DNA extraction protocol and PCR assay developed in this study can be used for rapid and reliable detection of L. maculans from infected seeds of oilseed rape .     

经皮穴位电刺激对老年髋关节置换术患者脑氧代谢以及术后认知功能、镇痛效果的影响

目的:探讨经皮穴位电刺激(TEAS)对老年髋关节置换术患者脑氧代谢以及术后认知功能、镇痛效果的影响。方法:选取2016年1月～2018年7月期间我院收治的行髋关节置换术患者91例为研究对象,将研究对象根据随机数字表法分为对照组(n=45)和研究组(n=46),对照组给予常规麻醉处理,研究组在对照组基础上给予TEAS,比较两组脑氧代谢、术后认知功能以及镇痛效果,记录两组术后的不良反应发生情况。结果:在降压开始后20 min(T1)、降压开始后40 min(T2)、停止降压后20 min(T3)时间点时,两组静脉血氧含量(Cjv O2)较降压前即刻(T0)时间点升高,且研究组高于对照组(P0.05),两组脑动-静脉血氧含量差(Da-jvO2)、脑氧摄取率(CERO2)较T0时间点降低,且研究组低于对照组(P0.05)。两组患者术后72h简易智能量表(MMSE)评分均较术前24h降低,但研究组高于对照组(P0.05),研究组术后认知功能障碍(POCD)发生率低于对照组(P0.05)。与术前比较,两组患者术后8h、术后24h、术后48h视觉疼痛模拟量表(VAS)评分均升高(P0.05),但研究组术后8h、术后24h、术后48h等时间点VAS评分均低于对照组(P0.05)。两组患者不良反应总发生率比较无差异(P0.05)。结论:TEAS对老年髋关节置换术后患者的镇痛效果确切,可有效改善脑氧代谢情况,提高术后认知功能,临床应用价值较高。     

经皮椎体后凸成形术对老年骨质疏松性椎体压缩骨折患者新发椎体骨折风险和生活质量的影响

目的:探讨经皮椎体后凸成形术(PKP)治疗老年骨质疏松性椎体压缩骨折的疗效及对患者新发椎体骨折风险和生活质量的影响。方法:选取2015年2月~2017年7月期间海南医学院第二附属医院收治的老年骨质疏松性椎体压缩骨折患者109例为研究对象,根据患者意愿和经济条件将其分为保守治疗组(n=54,采用非手术治疗)和PKP组(n=55,采用PKP治疗),比较两组患者临床指标、疼痛程度、椎体指标、生活质量、新发椎体骨折风险以及并发症发生情况。结果:PKP组住院时间、卧床时间低于保守治疗组(P0.05)。两组患者治疗1周后、治疗1个月后、治疗3个月后、治疗6个月后以及治疗12个月后视觉疼痛模拟评分(VAS)逐渐降低(P0.05),PKP组治疗1周后、治疗1个月后VAS评分低于保守治疗组(P0.05)。保守治疗组治疗12个月后椎体前缘高度丢失率、Cobb角均低于治疗前及治疗1周后(P0.05),PKP组治疗1周后、治疗12个月后椎体前缘高度丢失率、Cobb角逐渐降低(P0.05);PKP组治疗1周后、治疗12个月后椎体前缘高度丢失率、Cobb角低于保守治疗组(P0.05)。两组患者治疗12个月后标准生理组分(PCS)、标准心理组分(MCS)评分均较治疗前升高,且PKP组PCS、MCS评分高于保守治疗组(P0.05)。两组患者新发椎体骨折发生率比较差异无统计学意义(P0.05)。PKP组并发症发生率低于保守治疗组(P0.05)。结论:PKP治疗老年骨质疏松性椎体压缩骨折疗效确切,安全可靠,可缩短患者住院时间和卧床时间,缓解患者早期疼痛,改善患者生活质量和伤椎后凸畸形,且不会增加新发椎体骨折风险。