The usefulness of measuring n-butyric acid concentration as a new indicator of blood decomposition in forensic autopsy

In forensic medicine, although various alcohols have been reported as indicators of decomposition in collected blood, no studies have examined short-chain fatty acids as indicators. In this study, the blood n-butyric acid concentration was quantified, and the association between n-butyric acid and decomposition was investigated to determine whether the detection of n-butyric acid could be a new indicator of decomposition. Among the forensic autopsies performed from 2016 to 2018 in our laboratory, the cases were divided into decomposed (n = 20) and non-decomposed (n = 20) groups based on macroscopic findings. Blood samples collected at the time of autopsy were derivatized with 3-nitrophenylhydrazine hydrochloride after solid-phase extraction. The n-butyric acid concentration was measured using liquid chromatography–tandem mass spectrometry. In addition, ethanol and n-propanol were measured using a gas chromatography-flame ionization detector. There was a significant difference (p < 0.01) in the concentrations of n-butyric acid between the decomposed and non-decomposed groups (0.343 ± 0.259 [0.030–0.973] and 0.003 ± 0.002 [0.001–0.007] mg/mL, respectively). In the decomposed group, n-butyric acid was detected at high concentrations, even in cases where n-propanol was low. These results suggest that n-butyric acid is more likely to be an indicator of blood decomposition than n-propanol.     

Protective activities of ellagic acid and urolithins against kidney toxicity of environmental pollutants: A review

Oxidative stress and inflammation are two possible mechanisms related to nephrotoxicity caused by environmental pollutants. Ellagic acid, a powerful antioxidant phytochemical, may have great relevance in mitigating pollutant-induced nephrotoxicity and preventing the progression of kidney disease. This review discusses the latest findings on the protective effects of ellagic acid, its metabolic derivatives, the urolithins, against kidney toxicity caused by heavy metals, pesticides, mycotoxins, and organic air pollutants. We describe the chelating, antioxidant, anti-inflammatory, antifibrotic, antiautophagic, and antiapoptotic properties of ellagic acid to attenuate nephrotoxicity. Furthermore, we present the molecular targets and signaling pathways that are regulated by these antioxidants, and suggest some others that should be explored. Nevertheless, the number of reports is still limited to establish the efficacy of ellagic acid against kidney damage induced by environmental pollutants. Therefore, additional preclinical studies on this topic are required, as well as the development of well-designed clinical trials.     

Purification and characterization of the carbonic anhydrase enzyme from Black Sea trout (Salmo trutta Labrax Coruhensis) kidney and inhibition effects of some metal ions on enzyme activity

In this study, the carbonic anhydrase (CA) enzyme was purified from Black Sea trout (Salmo trutta Labrax Coruhensis) kidney with a specific activity of 603.77 EU/mg and a yield of 35.5% using Sepharose-4B-l-tyrosine- sulphanilamide affinity column chromatography. For determining the enzyme purity and subunit molecular mass, sodiumdodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) was performed and single band was observed. The molecular mass of subunit was found approximately 29.71 kDa. The optimum temperature, activation energy (E_a), activation enthalpy (ΔH) and Q₁₀ values were obtained from Arrhenius plot. K_m and V_max values for p-nitrophenyl acetate of the purified enzyme were calculated from Lineweaver-Burk graphs. In addition, the inhibitory effects of different heavy metal ions (Fe²⁺, Pb²⁺, Co²⁺, Ag⁺ and Cu²⁺) on Black Sea trout kidney tissue CA enzyme activities were investigated by using esterase method under in vitro conditions. The heavy metal concentrations inhibiting 50% of enzyme activity (IC₅₀) were obtained. Finally K_i values and inhibition types were calculated from Lineweaver-Burk graphs.     

Genetic variation of 17 autosomal STR loci in the Lahu population from Yunnan and phylogenetic structure exploration among 28 populations

This study evaluated the genetic variation of 17 autosomal short tandem repeat (STR) loci included in the PowerPlex® 18D Kit. Samples of 562 unrelated healthy Lahu individuals living in Yunnan Province in southwestern China were investigated. The data were analyzed to provide information on allele frequencies and other statistical parameters relevant to the forensic population. Of the 17 loci, 16 reached the Hardy–Weinberg equilibrium after Bonferroni correction. A total of 176 alleles were identified in 17 STR loci, and allele frequencies ranged from 0.000 890 to 0.578 292. The combined discrimination power (CPD) and probability of excluding paternity (CPE) of the 17 STR loci were 0.999 999 999 999 999 999 489 and 0.999 998 301 753 122. The genetic relationships among 28 populations were also estimated.     

力学刺激对3月龄骨质疏松大鼠成骨细胞增殖与合成功能的影响

探讨周期性双轴力学应变对成骨细胞增殖与分化合成功能的影响。将正常 3月龄雌性 SD大鼠和骨质疏松大鼠颅顶骨分离的成骨细胞分别在含 10 %胎牛血清的 F- 12培养液中培养 ,并接种在双轴力学应变装置中。当细胞生长至亚融合状态 (Subconfluence) ,给细胞施加力学刺激 ,频率为 1Hz,力学刺激分别为 4 0 0、10 0 0、4 0 0 0μ strain;作用时间分别为每天 30 m in,2、4、8h,共加载两天。以未受力学刺激的细胞为对照组 ,受力学刺激的细胞为实验组 ,并进行比较。采用流式细胞技术测定细胞增殖变化 ;采用同位素标记方法检测成骨细胞骨钙素、I型胶原 C端前肽 (PICP)和总蛋白的分泌量。结果表明 :1)在静态培养条件下 ,3ovx组与 3control组比较 ,其细胞功能活性无明显变化 ,但 3ovx组大鼠成骨细胞增殖活性明显增高 ,这与绝经后骨质疏松骨代谢的高转换率相一致。 2 )4 0 0、10 0 0 μstrain力学刺激可以促进 3control组成骨细胞 I型胶原、骨钙素和总蛋白的分泌量增加 ,促进成骨细胞的分化成熟 ;在 10 0 0μstrain力学刺激下 ,成骨细胞合成骨基质的能力增加最为明显。同时 ,在 4 0 0、10 0 0μstrain力学刺激的初期也可以促进成骨细胞的增殖 ,而促进成骨细胞的分化成熟的作用大于促进细胞增殖的作用。 3)在4 0 0 0 μ     

慢性束缚应激致大鼠持续性高血糖症与孤束核损伤有关

目的 探讨孤束核联合亚核前部（acNTS）损伤在慢性束缚应激（CRS）所致的胰岛素抵抗性高血糖症发生中的作用。 方法 采用CRS大鼠模型（n=20;7 d束缚+3 d自由活动，共40 d），检测葡萄糖代谢相关指标。 结果 CRS导致约1/3的个体（n=7）持续性的中度胰岛素抵抗性高血糖（空腹血糖不超过11 mmol/L）。CRS高血糖鼠acNTS可观察到神经元染色浓缩，Caspase-3表达和TUNEL阳性染色，提示出现神经元凋亡样改变。机械损伤acNTS（n=6），空腹血糖水平逐渐升高，也能导致胰岛素抵抗性高血糖, 且高胰岛素血症、胰岛平均体积增大和血清皮质酮水平不变等特点与CRS小鼠一致。 结论 CRS损伤了acNTS的葡萄糖敏感神经元，从而使血糖调节紊乱。     

应用复合诱导突变分离PCR法检测mtDNA的单核苷酸多态性

目的应用复合诱导突变分离PCR(multiplexed mutagenically separated PCR,MS-PCR)技术、银染分型,建立线粒体DNA(mitochondrial DNA,mtDNA)编码区单核苷酸多态(single nucleotide polymorphism,SNP)分型系统,探讨其应用价值。并调查了成都汉族群体mtDNA编码区4个SNP基因座等位基因频率和单倍型分布情况。方法根据SNP基因座(C12705T、A8701G、G8584A、C10400T)设计两条片段相差4个碱基的等位基因特异性引物和一条公共引物,4个SNP基因座复合扩增,PCR产物经聚丙烯酰胺凝胶电泳、银染显带后确定样本的基因型。结果不同SNP基因座为长度不同的单一谱带,其分型结果与直接测序一致。在成都汉族160名无关个体中,4个SNP基因座C12705T、A8701G、G8584A、C10400T等位基因频率分别为0.3813/0.6187、0.4813/0·5187、0.8250/0.1750、0.4938/0.5062;共检出6种单倍型,单倍型的基因多样性为0.7137。结论建立的MMS-PCR银染分型系统是一种简单、快速、准确、有效的SNP分型方法,对建立mtDNA编码区SNP数据库,研究群体遗传学、进化学和进行法医学个人识别和亲子鉴定有重要意义。     

黄背草与2种菅属植物叶绿体基因组特征比较及系统发育分析

目的 以黄背草Themeda japonica为试验材料,比较分析其与阿拉伯黄背草T. triandra、中华菅T. quadrivalvis2种同属植物的叶绿体基因组特征及其与近缘物种的系统发育关系。方法 采用Illumina HiSeq高通量测序平台首次对黄背草叶绿体基因组进行测序,使用SPAdes和CpGAVAS2分别对其进行组装和注释,并用Codon W、DnaSP和MISA等对其与2种同属植物进行一系列比较基因组分析,利用最大似然法（maximum likelihood,ML）构建系统进化树。结果 3个叶绿体基因组全长为138 735～138 961 bp,具有典型的四分体结构,共注释出129个基因;黄背草与其同属的2个物种相比,反向重复区（inverted repeats,IR）收缩了2132 bp,大单拷贝区（large single copy region,LSC）扩张了约4000 bp,而小单拷贝区（small single copy region,SSC）变化不大。密码子偏好性分析显示,3个叶绿体基因组相对丰度最大和最小的密码子都相同,同义密码子相对使用丰度略有不同...     

miR-146a-5p regulates autophagy and NLRP3 inflammasome activation in epithelial barrier damage in the in vitro cell model of ulcerative colitis through the RNF8/Notch1/mTORC1 pathway

Ulcerative colitis (UC) is a chronic inflammatory disease affecting the colon that can be influenced by microRNAs (miRNAs). This study aims to investigate the impact of miR-146a-5p on lipopolysaccharide (LPS)-induced Caco-2/HT-29 cell autophagy and NLRP3 inflammasome activation and the underlying mechanism, with the aim of identifying potential therapeutic targets. We used LPS to establish Caco-2/HT-29 cell models and measured cell viability by CCK-8. The levels of miR-146a-5p, RNF8, markers of NLRP3 inflammasome activation and autophagy, proteins involved in the Notch1/mTORC1 pathway, and inflammatory factors were assessed by RT-qPCR, Western blot, and ELISA. Intestinal epithelial barrier function was evaluated by measuring transepithelial electrical resistance. Autophagic flux was measured using tandem fluorescent-labeled LC3. miR-146a-5p was highly-expressed in LPS-induced Caco-2/HT-29 cells, and autophagy flux was blocked at the autolysosomal stage after LPS induction. Inhibition of miR-146a-5p suppressed NLRP3 inflammasome activation, reduced intestinal epithelial barrier damage, and facilitated autophagy inhibition in LPS-induced Caco-2/HT-29 cells. The autophagy inhibitor NH4Cl partially nullified the inhibitory effects of miR-146a-5p inhibition on NLRP3 inflammation activation. miR-146a-5p targeted RNF8, and silencing RNF8 partly abrogated the action of miR-146a-5p inhibition on promoting autophagy and inhibiting NLRP3 inflammasome activation. miR-146a-5p inhibition suppressed the Notch1/mTORC1 pathway activation by upregulating RNF8. Inhibition of the Notch1/mTORC1 pathway partially nullified the function of silencing RNF8 on inhibiting autophagy and bolstering NLRP3 inflammasome activation. In conclusion, miR-146a-5p inhibition may be a potential therapeutic approach for UC, as it facilitates autophagy of LPS-stimulated Caco-2/HT-29 cells, inhibits NLRP3 inflammasome activation, and reduces intestinal epithelial barrier damage by upregulating RNF8 and suppressing the Notch1/mTORC1 pathway.     

间充质祖细胞源性的神经元样细胞肌内移植后自身特性的维持

目的探讨间充质祖细胞(mesenchymal progenitor cells,MPCs)源性的神经元样细胞肌内移植后能否维持其自身特性。方法取3周龄健康GFP转基因C57小鼠后肢长骨进行MPC培养及鉴定,选取生长良好的第3代MPC,采用神经元原代培养上清液进行神经元样细胞诱导分化后收集细胞,并制备成5×105/μL细胞悬液备用。选取12周龄健康C57小鼠24只,按随机数字表法分为损伤+移植细胞组、损伤+生理盐水组、假手术+移植细胞组、假手术+生理盐水组。损伤+移植细胞组及假手术+移植细胞组:将制备的5μL MPC悬液散点注入小鼠三头肌内;损伤+生理盐水组和假手术+生理盐水组:同法注入等量生理盐水。术后4周取材,荧光显微镜下观察肌内移植细胞存活情况,电镜观察肌肉超微结构的变化,免疫荧光染色检测神经元特异性标志物NSE、NeuN及神经胶质特异性标志物GFAP的表达情况。结果肌内移植细胞生长良好且均匀分布于肌细胞间隙,并抑制了肌细胞核、线粒体、内质网的退变及肌肉纤维化的发生。损伤+移植细胞组定植存活的移植细胞阳性表达:NSE(58.35±1.37)%、NeuN(60.22±0.16)%及GFAP(13.32±1.65)%,损伤+生理盐水组、假手术+移植细胞组、假手术+生理盐水组均无阳性细胞表达。结论 MPC具有良好的生物学活性,经体外诱导分化为神经元及神经胶质样细胞后移植于失神经支配肌内可定植存活,并能够维持神经元及神经胶质样细胞的特性,同时有效抑制了肌细胞核、线粒体、内质网的退变及肌肉纤维化的发生。