不同预处理对小鼠粪样菌群高通量测序分析结果的影响

应用高通量测序技术比较不同预处理对小鼠粪样菌群结构的影响,以期为后续相关研究提供参考依据。采集昆明小鼠新鲜粪样,分为原始粪样组、生理盐水处理组和PBS处理组,提取粪样DNA,采用Illumina MiSeq平台进行测序对3组样本的16S rRNA V3~V4区基因文库进行生物信息学分析。结果显示,3组样本拥有共同的OTUs 188个,不同预处理对粪样微生物多样性和丰度产生影响,生理盐水处理组、PBS处理组和原始粪样组分别检出14、11、12个门,3组样本共有门11个;分别检出24、21、21个纲,3组样本共有纲20个;分别检出62、55、59个科,3组样本共有科26个;分别检出147、126、137个属,3组样本共有属117个。总体来看,生理盐水处理组微生物多样性和丰度相对最高,原始粪样组次之,PBS处理组最低。粪样经生理盐水处理后,能在一定程度上提高肠道菌群检测的准确性。     

Detection of a new 20-bp insertion/deletion (indel) within sheep PRND gene using mathematical expectation (ME) method

Prion-related protein doppel gene (PRND), as an essential member of the mammalian prion gene family, is associated with the scrapie susceptibility as well as phenotype traits, so the genetic variation of the PRND has been highly concerned recently, including the single nucleiotide polymorphism (SNP) and insertion/deletion (indel). Therefore, the objective of present study was to examine the possible indel variants by mathematical expectation (ME) detection method as well as explore its associations with phenotype traits. A novel 20-bp indel was verified in 623 tested individuals representing 4 diversity sheep breeds. The results showed that 3 genotypes were detected and the minor allelic frequency were 0.008 (Lanzhou Fat-Tail sheep, LFTS), 0.084 (Small Tail Han sheep, STHS), 0.021(Tong sheep, TS) and 0.083 (Hu sheep, HS), respectively. Comparing with the traditional method of detecting samples one by one, the reaction times with ME method was decreased by 36.22% (STHS), 37.00% (HS), 68.67% (TS) and 83.33% (LFTS), respectively. Besides, this locus was significantly associated to cannon circumference index (P = 0.012) and trunk index (P = 0.037) in the Hu sheep breed. Notably, it was not concordance with the present result of DNA sequencing (GCTGTCCCTGCAGGGCTTCT) and dbSNPase of NCBI (NC_443194: g.46184887- 46184906delCTGCTGTCCCTGCAGGGCTT). Consequently, it was the first time to detect the new 20-bp indel of sheep PRND gene by ME strategy, which might provide a valuable theoretical basis for marker-assisted selection in sheep genetics and breeding.     

20(S)-Protopanaxadiol Inhibition of Progression and Growth of Castration-Resistant Prostate Cancer

Castration-resistant progression of prostate cancer after androgen deprivation therapies remains the most critical challenge in the clinical management of prostate cancer. Resurgent androgen receptor (AR) activity is an established driver of castration-resistant progression, and upregulation of the full-length AR (AR-FL) and constitutively-active AR splice variants (AR-Vs) has been implicated to contribute to the resurgent AR activity. We reported previously that ginsenoside 20(S)-protopanaxadiol-aglycone (PPD) can reduce the abundance of both AR-FL and AR-Vs. In the present study, we further showed that the effect of PPD on AR expression and target genes was independent of androgen. PPD treatment resulted in a suppression of ligand-independent AR transactivation. Moreover, PPD delayed castration-resistant regrowth of LNCaP xenograft tumors after androgen deprivation and inhibited the growth of castration-resistant 22Rv1 xenograft tumors with endogenous expression of AR-FL and AR-Vs. This was accompanied by a decline in serum prostate-specific antigen levels as well as a decrease in AR levels and mitoses in the tumors. Notably, the 22Rv1 xenograft tumors were resistant to growth inhibition by the next-generation anti-androgen enzalutamide. The present study represents the first to show the preclinical efficacy of PPD in inhibiting castration-resistant progression and growth of prostate cancer. The findings provide a rationale for further developing PPD or its analogues for prostate cancer therapy.     

Screening for cold-resistant tomato under radiation mutagenesis and observation of the submicroscopic structure

Tomato is a cold-sensitive crop that is vulnerable to chilling injury. To screen for chilling resistance in tomato mutants, seeds were irradiated with different doses of ⁶⁰Co-γ and then evaluated according to the five stages of the chilling injury index. Moreover, physiological indexes and observation of the submicroscopic structure by electron microscopy were used to examine cold resistance in the mutants. The physiological index results showed much higher cold resistance in the mutants compared to wild type. The cellular structures of the cold-resistant mutants were more complete than in wild-type plants after chilling treatment. The chloroplasts of the mutants were close to the cell periphery; the double membrane structure of the chloroplast was intact, and well-developed granal stacks were interconnected by stroma. Compared to wild-type plants, the stomatal density of cold-resistant mutants increased considerably, though the stomatal size showed no obvious changes.     

Dexmedetomidine Protects against Transient Global Cerebral Ischemia/Reperfusion Induced Oxidative Stress and Inflammation in Diabetic Rats

Background

Transient global cerebral ischemia/reperfusion (I/R) is a major perioperative complication, and diabetes increases the response of oxidative stress and inflammation induced by I/R. The objective of this study was to determine the protective effect of dexmedetomidine against transient global cerebral ischemia/reperfusion induced oxidative stress and inflammation in diabetic rats.

Methods

Sixty-four rats were assigned into four experimental groups: normoglycemia, normoglycemia + dexmedetomidine, hyperglycemia, and hyperglycemia + dexmedetomidine and all subsequent neurological examinations were evaluated by a blinded observer. Damage to the brain was histologically assessed using the TUNEL staining method while western blotting was used to investigate changes in the expression levels of apoptosis-related proteins as well as the microglia marker, ionized calcium-binding adapter molecule 1 (Iba1). Water content in the brain was also analyzed. In addition, hippocampal concentrations of malondialdehyde (MDA) and Nox2 (a member of the Nox family of NADPH oxidases), and the activity of superoxide dismutase and catalase were analyzed. Finally, changes in serum concentrations of tumor necrosis factor-α (TNF-α) and interleukin (IL)-6 were detected.

Results

Results showed that diabetes increased brain water content, the number of apoptotic neurons, early neurological deficit scores, oxidative stress (MDA and Nox2) and inflammation (pro-inflammatory cytokines including TNF-α and IL-6) levels following transient global I/R injury, but that these symptoms were attenuated following administration of dexmedetomidine.

Conclusions

These findings suggest that dexmedetomidine can significantly alleviate damage resulting from I/R, and this mechanism may be related to a reduction in both oxidative stress and inflammation which is normally associated with I/R.     

Autophagy protects PC12 cells against deoxynivalenol toxicity via the Class III PI3K/beclin 1/Bcl-2 pathway

Deoxynivalenol (DON) is a major mycotoxin from the trichothecene family of mycotoxins produced by Fusarium fungi. It can cause a variety of adverse effects on human and farm animal health. Here, we determined the effect of DON on the Class III phosphatidylinositol 3-kinase (PIK3C3)/beclin 1/B cell lymphoma-2 (Bcl-2) pathway in PC12 cells and the relationship between autophagy and apoptosis. The effects of DON were evaluated based on the apoptosis ratio; the typical indicators of autophagy, including cellular morphology, acridine orange- and monodansylcadaverine-labeled vacuoles, green fluorescent protein–microtubule associated protein 1 light chain 3 (LC3) localization, and LC3 immunofluorescence; and the expression of key autophagy-related genes and proteins, that is, PIK3C3, beclin 1, Bcl-2, LC3, and p62. The relationship between autophagy and apoptosis was analyzed by western blot analysis and flow cytometry. DON-induced PC12 cell morphological changes and autophagy significantly. PIK3C3, beclin 1, and LC3 increased in tandem with the DON concentration used; Bcl-2 and p62 expression decreased as DON concentrations increased. Moreover, the PIK3C3/beclin 1/Bcl-2 signaling pathway played a role in DON-induced autophagy. Our findings suggest that DON can induce autophagy by activating the PIK3C3/beclin 1/Bcl-2 signaling pathway and that autophagy may play a positive role in reducing DON-induced apoptosis.     

Novel 3-dimensional dendrimer platform for glycolipid microarray

Glycolipids are important biological molecules that modulate cellular recognitions and pathogen adhesions. In this paper, we report a sensitive glycolipid microarray for non-covalently immobilizing glycolipids on a microarray substrate and we perform a set of immunoassays to explore glycolipid-protein interactions. This substrate utilizes a three-dimensional hydrazide-functionalized dendrimer monolayer attached onto a microscopic glass surface, which possesses the characteristics to adsorb glycoliplids non-covalently and facilitates multivalent attributes on the substrate surface. In the proof-of-concept experiments, gangliosides such as GM1, FucGM1, GM3, GD1b, GT1b, and GQ1b, and a lipoarabinomannan were tested on the substrate and interrogated with toxins and antibodies. The resulting glycolipid microarrays exhibited hypersensitivity and specificity for detection of glycolipid-protein interactions. In particular, a robust and specific binding of a pentameric cholera toxin B subunit to the GM1 glycolipid spotted on the array has demonstrated its superiority in sensitivity and specificity. In addition, this glycolipid microarray substrate was used to detect lipoarabinomannan in buffer within a limit-of-detection of 125 ng/mL. Furthermore, Mycobacterium tuberculosis (Mtb) Lipoarabinomannan was tested in human urine specimens on this platform, which can effectively identify urine samples either infected or not infected with Mtb. The results of this work suggest the possibility of using this glycolipid microarray platform to fabricate glycoconjugate microarrays, which includes free glycans and glycolipids and potential application in detection of pathogen and toxin.