应用iTRAQ-LC-MS/MS方法筛选大鼠DAI后脑组织差异表达蛋白质

目的应用相对和绝对定量同位素标记结合液相色谱-串联质谱法(isobaric tag for relative and absolute quantification-liquid chromatograph-mass spectrometer/mass spectrometer,iTRAQ-LC-MS/MS)筛选SD大鼠弥漫性轴索损伤(diffuse axonal injury,DAI)后脑组织差异表达的蛋白质,寻找诊断DAI的潜在生物标志物。方法参照Marmarou法建立DAI动物模型,分为空白对照组(n=4)、假打击组(n=4)和打击致死组(n=4),采用iTRAQ-LC-MS/MS技术对大鼠脑组织蛋白质进行检测,并对检测结果进行生物信息学分析及验证,筛选出差异表达的蛋白质。结果定量检测出2016种蛋白质,生物信息学分析显示其在细胞中分布广泛、功能多样,并且参与了多种生物过程,16种蛋白质在打击致死组具有差异表达,包括1种表达上调蛋白质和15种下调蛋白质;Western印迹法验证了iTRAQ-LC-MS/MS的结果。结论 iTRAQ-LC-MS/MS的检测技术筛选出大鼠DAI后脑组织中多个差异表达的蛋白质,不仅为深入探讨DAI后轴索损伤的发病机制提供了研究方向,也为DAI的诊断提供了潜在生物标志物。     

MALDI-TOF-IMS分析DAI大鼠脑组织内差异蛋白的分布

目的基于基质辅助激光解吸/电离-飞行时间质谱成像(matrix assisted laser desorption/ionizationtime of flight imaging mass spectrometry,MALDI-TOF-IMS)技术建立弥漫性轴索损伤(diffuse axonal injury,DAI)大鼠脑组织内蛋白差异化表达的质谱成像方法。方法应用AutoflexⅢ MALDI-TOF质谱仪在质荷比1 000~20 000范围内对DAI组和对照组脑组织进行质谱扫描,利用Clin Pro Tool 2.2软件对两组数据进行统计学分析,挑选出存在表达差异的蛋白质分子进行质谱成像,观察不同质荷比的蛋白质分子在脑组织中的分布情况。结果质荷比为4 963、5 634、6 253、6 714及7 532的5个蛋白质分子在DAI大鼠脑组织内表达量存在差异。结论 MALDI-TOF-IMS可用于大鼠DAI后脑组织内差异蛋白的研究,初步建立了质谱成像技术检测DAI大鼠脑组织中差异蛋白分布的分析方法。     

血清蛋白芯片表达谱在DVT疾病法医鉴定中的应用研究

目的利用蛋白芯片技术获取人体和大鼠深静脉血栓(deep venous thrombosis, DVT)组与对照组血清蛋白表达谱的蛋白峰差异,探讨蛋白表达谱与深静脉血栓形成的相关性,为DVT临床和法医学诊断提供依据。方法采用蛋白芯片技术检测DVT患者和正常人(n=10)的血清样本,获得血清蛋白芯片表达谱。建立标准化SD大鼠下腔静脉血栓模型,大鼠被随机分为DVT组和对照组(n=10),蛋白芯片技术检测获得各组血清蛋白表达谱;采用SIMCA-P 14.1软件的正交偏最小二乘判别(orthogonal partial least square-discriminant analysis,OPLS-DA)模型和Python软件的随机森林(Random forest, RF)算法模型对人体和大鼠数据进行判别分析。结果 DVT患者和正常人血清蛋白表达峰存在差异,OPLS-DA模型对DVT组和对照组准确判别,R~2X=0.631,R~2Y=0.928,Q~2=0.796;动物模型DVT组与对照组的血清蛋白表达峰亦存在差异,OPLS-DA模型可以将DVT组和对照组准确判别,R~2X=0.735,R~2Y=0.953,Q~2=0.900;人体和动物模型血清中共有8个分子量相似的血清蛋白峰,RF算法模型准确率为100%,人体预测准确率为65%。结论蛋白芯片技术可以快速获取DVT人体和大鼠血清蛋白表达谱,建立OPLS-DA模型和RF模型能对DVT组和对照组进行分类预测分析,为DVT疾病诊断和法医学鉴定提供新的辅助方法。     

心肌缺血早期表达蛋白的质谱研究

目的用SELDI-TOFMS观察大鼠急性心肌缺血后心肌蛋白质谱的变化规律,筛选心肌缺血的蛋白标记物,为法医学心源性猝死的诊断提供依据。方法SD大鼠分成手术组,假手术组和非手术对照组。在结扎大鼠冠脉前降支后5min～6h内不同的时间段取大鼠心肌组织。蛋白提取后用CM10芯片结合SELDI-TOFMS质谱仪检测。结果血清CM10芯片检测到3个缺血后蛋白峰,质荷比(m/z)分别为6304Da、8337Da、8375Da,在手术组中阳性率和特异性均为100%,假手术组和非手术组中均没有发现。特异性升高的峰有3个:峰6658Da、6876Da,都是在实验组中≥15min以后升高2倍以上,8577Da峰则在缺血≥5min组升高。结论大鼠心肌组织中6304Da、8337Da、8375Da蛋白峰都是急性心肌缺血后的较为特异性改变,可在心肌缺血后5min内出现,有可能做为心肌缺血的蛋白标记,亦可能用于心肌缺血和法医学心源性猝死的诊断。     

GYPA、CD68和TNF-α蛋白鉴别水中晚期尸体生前死后伤的作用研究

目的 探讨血型糖蛋白A(Glycophorin A, GYPA)、CD68和肿瘤坏死因子-α(Tumor necrosis factor-α,TNF-α)蛋白在水中晚期尸体生前伤和死后伤的表达情况,为在水中晚期生前死后伤鉴定筛选标志物。方法 18只SD大鼠分为对照组（n=6）、生前挫伤浸没组（n=6）和死后挫伤浸没组（n=6）,生前挫伤浸没组采用机械失重技术建立SD大鼠右后肢挫伤模型,死后挫伤浸没组是SD大鼠处死后用机械失重技术建立右后肢挫伤模型,对照组不予致伤处理,所有大鼠均浸没72 h。肉眼观察及HE染色观察生前伤和死后伤的情况;免疫组织化学染色（Immunohistochemistry,IHC）和蛋白质免疫印迹（Western blot）检测GYPA、CD68及TNF-α蛋白在挫伤组织表达情况;随后在实际案例中予以验证。结果 肉眼观察和HE染色均无法较好地鉴别浸没72 h大鼠的生前伤和死后伤。免疫组织化学染色发现生前挫伤浸没组软组织GYPA表达呈阳性,CD68在炎性细胞膜呈阳性表达,TNF-α在胞质表达呈弱阳性,GYPA、CD68和TNF-α蛋白表达均显著高于死后挫伤浸没组和对照...     

二维红外光谱用于急性心肌缺血猝死的鉴别研究

目的采用衰减全反射-傅里叶变换红外光谱(ATR-FTIR)技术检测急性心肌缺血猝死大鼠的血清分子特征,探讨二维红外光谱鉴别急性心肌缺血猝死的法医学价值。方法成年雄性SD大鼠随机分为急性心肌缺血猝死组(实验组)和空气栓塞死组(对照组),血清样本行ATR-FTIR检测,应用OMNIC软件进行光谱预处理并求二阶导数光谱,分别计算两组原始光谱及二阶导数光谱的相似度,并应用Matlab R2010a软件建立二维光谱模型。结果实验组与对照组的大鼠血清的原始红外光谱在峰形、峰位、峰强度均极其相似,相似度达99.78%,二阶导数光谱显示出两者相似度降至98.62%,二维光谱模型显示两组光谱在1 625cm~(-1)、1 550cm~(-1)、1 080cm~(-1)及860cm~(-1)附近的成分差异,且它们的自动峰个数不相同,提示两组在蛋白质、糖原、核酸等分子含量存在较明显差异。结论仅从一维光谱上难以区分出两组的分子差异,二维红外光谱可以直观、有效地反映出急性心肌缺血猝死与空气栓塞死的大鼠血清分子差异,有望应用于法医学死因鉴别。     

大鼠慢性心衰致死的代谢特征与标记物研究

目的探寻慢性心衰(CHF)致死的相关代谢特征与潜在诊断标记物。方法阿霉素腹腔注射建立CHF大鼠模型,根据期间是否发生死亡分为心衰死亡(HFD)及存活组(HFS)。应用GC-MS检测心肌代谢谱,结合代谢组学策略分析HFD与HFS大鼠心肌代谢轮廓差异,筛选与CHF死亡相关代谢特征与潜在诊断标记物。结果制备了50例CHF大鼠,22例死亡,模型大鼠LVSP与±dp/dt max低于正常水平(p0.05)。正交偏最小二乘法-判别分析(OPLS-DA)得分图提示HFD与HFS大鼠心肌代谢特征差异显著;与HFS相比,HFD心肌的丁酸、尿素、亮氨酸、异亮氨酸、苯丙氨酸、鸟氨酸、嘌呤、赖氨酸上调;乳酸、脯氨酸、半胱氨酸、谷氨酰胺下调;反映了死亡大鼠心肌能量代谢、氨基酸代谢、鸟氨酸循环等通路受显著扰动。此外,苯丙氨酸、亮氨酸有较高的区分HFD与HFS的准确率(AUC、敏感性、特异性分别为:0.84,84.5%,80.9%;0.98,95.2%,95.7%)。结论 CHF致死大鼠心肌较存活大鼠代谢显著改变;苯丙氨酸、亮氨酸有较高的鉴别CHF死亡与存活大鼠的价值,有望为法医鉴定及临床防治提供参考。     

基于代谢组学技术的毒鼠强中毒机制研究

目的 研究毒鼠强中毒后大鼠的血浆代谢组学随时间变化规律，从代谢组学角度揭示其毒理学作用机制，为毒鼠强中毒的相关法医学研究提供依据。方法 将SD大鼠随机分成对照组和实验组(n=8)，实验组大鼠灌胃1/4LD₅₀(0.05 mg/kg)毒鼠强乙酸乙酯溶液，对照组灌胃等剂量乙酸乙酯溶液，灌胃后于24 h、5 d、16 d眼眶静脉采血(约1 mL)，应用超高效液相色谱-飞行时间质谱联用技术检测血浆中的内源性小分子物质。采用主成分分析(PCA)、正交偏最小二乘判别分析(OPLS-DA)等筛选显著差异代谢物，寻找与毒鼠强中毒相关的代谢通路。结果本研究以变量权重重要性排序(VIP)值>1和student t检验的P值<0.05为筛选标准，中毒后不同时间点共筛选出显著差异代谢物30种，包括鞘脂类、氨基酸类和脂肪酸类化合物，涉及到鞘脂信号通路、鞘脂代谢和ABC蛋白转运等10条代谢通路。结论 毒鼠强中毒后机体多条代谢通路受到显著影响，主要是不同程度的神经系统调节紊乱和能量代谢紊乱，整体变化呈现出累积效应，可为研究毒鼠强毒性作用机制提供依据。     

PCR-DGGE法检测浮游生物16S rDNA在溺死鉴定中的应用

目的建立并评估PCR-DGGE法检测浮游生物对溺死鉴定的应用价值。方法大白兔30只随机分为3组:溺死组(n=12),死后抛尸组(n=12)和对照组(n=6);溺死组和死后抛尸组又分为2个亚组:东湖水域组(n=6)和墨水湖水域组(n=6)。死后提取心血和肺、肝、肾、脑等组织,匀浆后,采用Percoll密度梯度离心法分离浮游生物并提取其DNA,PCR扩增浮游生物特异的16S rDNA片段后分别用琼脂糖凝胶电泳及DGGE检测分析。2个溺死案例检材同法检验。结果溺死组各组织器官中浮游生物检测多呈阳性:肺(100%)、肝(83%)、肾(75%)、心血(83%)、脑(42%);死后抛尸组仅2例肺组织(16.7%)检出阳性;对照组全部阴性。溺尸肺组织DGGE分型图谱与相应溺死点水样分型图谱相似,而与非溺死点水样分型结果差异显著。2实际案例均呈阳性。结论本方法不仅有助于定性诊断溺死,而且通过比较产物的多样性可以推断溺死地点,在法医学溺死鉴定中具有较大实用价值。     

CT、MRI在弥漫性轴索损伤法医学鉴定中应用价值分析

目的探讨CT和MRI检查方法在弥漫性轴索损伤(diffuse axonal injury,DAI)法医学鉴定中的应用价值。方法收集26例CT和MRI影像资料齐全的DAI鉴定资料进行回顾性分析,比较CT和MRI两种方法对DAI病灶检出率的差异,分析DAI病灶检出数量与法医学鉴定结果之间的关系。结果 26例经MRI检出DAI病灶787个,经CT检出DAI病灶32个,MRI对DAI病灶的敏感度显著高于CT(P0.01)具有统计学意义。26例DAI鉴定的伤残等级数值与影像学检查发现的病灶数量之间存在负相关(r=-0.908,P0.01)。结论 MRI检查对DAI病灶的检出率较CT检查具有明显优势,可为DAI诊断提供更为准确的依据,在法医临床学鉴定中具有重要的应用价值。     

Bhartṛhari and Maṇḍana on Avidyā

The concept of avidyā is one of the central categories in the Advaita of Śaṇkara and Maṇḍana. Shifting the focus from māyā, interpreted either as illusion or as the divine power, this concept brings ignorance to the forefront in describing duality and bondage. Although all Advaitins accept avidyā as a category, its scope and nature is interpreted in multiple ways. Key elements in Maṇḍana’s philosophy include the plurality of avidyā, individual selves as its substrate and the Brahman as its field (viṣaya), and the distinction in avidyā between non-apprehension and misapprehension. A closer investigation shows that Maṇḍana is directly influenced by Bhartṛhari’s linguistic non-dualism in developing the concept of avidyā. This study also compares other key constituents such as vivartta and pariṇāma that are relevant to the analysis of avidyā. As the concept of counter-image (pratibimba) emerges as a distinct stream of Advaita subsequent to Maṇḍana, this study also compares the application of pratibimba in the writings of Bhartṛhari and Maṇḍana.     

Unraveling the <Emphasis Type="Italic">Kāvyaprakāśa</Emphasis>: Jayadeva Pīyūṣavarṣa’s idiosyncratic sequence of topics in the <Emphasis Type="Italic">Candrāloka</Emphasis>

In his twelfth century alaṃkāraśāstra, the Candrāloka, Jayadeva Pīyūṣavarṣa reverses the sequence of topics found in Mammaṭa’s Kāvyapr-akāśa, an earlier and immensely popular work. With such a structural revisionism, Jayadeva asserts the autonomy of his own work and puts forth an ambitious critique of earlier approaches to literary analysis. Jayadeva investigates the technical and aesthetic components of poetry in the first part of the Candrāloka, prior to his formal semantic investigations in the latter half of the text, thus suggesting that aesthetic evaluations of poetry beneficially inform scientific investigations of language. Jayadeva’s organization of his chapters on the semantic operations, moreover, intimates that the study of suggestive and metaphoric functions of language clarifies our understanding of denotation, which is conventionally understood to be the primary and direct path of verbal designation.     

The Supreme Array Scripture: A New Interpretation of the Title “Gaṇḍavyūha-sūtra”

This article argues for a new interpretation of the Sanskrit compound gaṇḍa-vyūha as it is used in the common title of the Mahāyāna text the Gaṇḍavyūha-Sūtra.The author begins by providing a brief history of the sūtra’s appellations in Chinese and Tibetan sources. Next, the meanings of gaṇḍa (the problematic member of the compound) are explored. The author proposes that contemporary scholars have overlooked a meaning of gaṇḍa occurring in some compounds, wherein gaṇḍa can mean simply “great,” “big” or “massive.” This general sense is particularly common in the compound gaṇda-śaila (a “massive rock” or “boulder”) and is found in such texts as the Bhāgavata Purāṇa, the Harivaṃśa and the Harṣacarita. Following the discussion of Gaṇḍa, the author examines the term vyūha (“array”) as it is used in the Gaṇḍavyūha-sūtra. The article concludes with the suggestion that a more appropriate translation of the Gaṇḍavyūha-sūtra would be “The Supreme array Scripture.”     

Evaluation of six presumptive tests for blood, their specificity, sensitivity, and effect on high molecular-weight DNA

Luminol, leuchomalachite green, phenolphthalein, Hemastix, Hemident, and Bluestar are all used as presumptive tests for blood. In this study, the tests were subjected to dilute blood (from 1:10,000 to 1:10,000,000), many common household substance, and chemicals. Samples were tested for DNA to determine whether the presumptive tests damaged or destroyed DNA. The DNA loci tested were D2S1338 and D19S433. Leuchomalachite green had a sensitivity of 1:10,000, while the remaining tests were able to detect blood to a dilution of 1:100,000. Substances tested include saliva, semen, potato, tomato, tomato sauce, tomato sauce with meat, red onion, red kidney bean, horseradish, 0.1 M ascorbic acid, 5% bleach, 10% cupric sulfate, 10% ferric sulfate, and 10% nickel chloride. Of all the substances tested, not one of the household items reacted with every test; however, the chemicals did. DNA was recovered and amplified from luminol, phenolphthalein, Hemastix, and Bluestar, but not from leuchomalachite green or Hemident.     

Monks Who Have Sex: Pārājika Penance in Indian Buddhist Monasticisms

In the study of Buddhism it is commonly accepted that a monk or nun who commits a pārājika offence is permanently and irrevocably expelled from the Buddhist monastic order. This view is based primarily on readings of the Pāli Vinaya. With the exception of the Pāli Vinaya, however, all other extant Buddhist monastic law codes (Dharmaguptaka, Mahāsāṅghika, Mahīśāsaka, Sarvāstivāda and Mūlasarvāstivāda) contain detailed provisions for monks and nuns who commit pārājikas but nevertheless wish to remain within the saṅgha. These monastics are not expelled. Rather, they are granted a special status known as the śikṣādattaka. In this paper I explore the rules. concerning pārājika penance and the śikṣādattaka with specific regard to monastic celibacy. Given that five out of six extant law codes recognise this remarkable accommodation to the rule of celibacy, I argue that we must look to Vinayas other than the Pāli Vinaya if we are to arrive at a nuanced and representative view of Indian Buddhist monasticism.

Grant me chastity and continence, but not yet. Augustine of Hippo (354–430 C.E.)

     

Family breakdown and the ‘Welfare Child’ in 19th and 20th century Britain

This article traces new cycles of interest in past children as distinct from past childhood. Recent work highlighting that a conceptualisation of childhood existed even in periods with few written records closes the chapter begun by Philippe Ariès in 1960. Instead, there has been a new surge of interest in children on the edges of family life, as well as children in similarly liminal positions between the worlds of adults and children: runaways, delinquents and orphans. Several themes in the literature are identified, based on the conflicting ideas of ‘body/mind’, ‘victim/threat’, ‘needs/rights’. It is noted that researchers are using more imaginative ways of reaching the lived experience of children than the family or institutional framework, and that an increasing link is drawn between historical and modern concerns such as child abuse and the care of ‘at risk’ children.     

Vedic Language and Vaiṣṇava Theology:Madhva’s Use of Nirukta in his Ṛgbhāṣya

This article explores the way in which Madhva (1238–1317), the founder of the Dvaita Vedānta system of Hindu thought, reformulates the traditional exegetic practice of nirukta or “word derivation” to validate his pluralistic, hierarchical, and Vaiṣṇava reading of the Ṛgvedic hymns. Madhva’s Ṛgbhāṣya (RB) is conspicuous for its heavy reliance on and unique deployment of this exegetical tactic to validate several key features of his distinctive theology. These features include his belief in Viṣṇu’s unique possession of all perfect attributes (guṇaparipūrṇatva) and His related conveyability by all Vedic words (sarvaśabdavācyatva). Such an understanding of Vedic language invokes the basic nirukta presupposition that words are eternally affiliated with the meanings they convey. But it is also based onMadhva’s access to a lexicon entitled Vyāsa’s Nirukti with which his critics and perhaps even his commentators seem to be unfamiliar.While the precise status of this text is the subject of ongoing debate, Madhva’s possession of special insight into the sacred canon is established in part by his unique claim to be an avatāra of the wind god Vāyu and a direct disciple of Viṣṇu Himself in the form of Vyāsa₁. Thus, Madhva’s use of nirukta invokes his personal charisma to challenge not only conventional understandings of the hymns but traditional exegetic norms. Madhva’s provision of an alternative tradition of nirukta provoked sectarian debate throughout the Vijayanagara period over the extent to which one could innovate in established practices of reading the Veda. Articulating the Veda’s precise authority was a key feature of Brahmin debates during this period and reflects both the empire’s concern with promoting a shared religious ideology and the competition among rival Brahman sects for imperial patronage that this concern elicited. By looking at how two of Madhva’s most important commentators (the 14^th-century Jayatīrtha and the 17^th-century Rāghavendra) sought to defend his niruktis, this article will explore how notions of normative nirukta were articulated in response to Madhva’s deviations. At the same time, however, examining Madhva’s commentators’ defense of his niruktis also demonstrates the extent to which Madhva actually adhered to selected exegetic norms. This reveals that discomfort with Madhva’s particular methods for deriving words stemmed, in part, from a more general ambivalence towards this exegetical tactic whose inherent open-endedness threatened to undermine the fixity of the canon’s very substance: its language. Vyāsa’s Nirukti is one of several ”unknown sources” cited in Madhva’s commentaries whose exact status continues to be debated. Some scholars (e.g. Rao, Sharma, Siauve) maintain that these texts are part of a now lost Pāṅcarātra tradition that Madhva is attempting to preserve. This may be true for many of these citations. However, in addition to claiming to be both an avatāra of Vāyu and Viṣṇu-as-Vyāsa’s student, Madhva states in several places (e.g., VTN 42, RB 162) that the canon has suffered loss during transmission and that only Viṣṇu can reveal it in its entirety. Thus, it is possible that Madhva intends texts like Vyāsa’s Nirukti to be viewed as part of an ongoing and corrective revelation, a notion that is compatible with many Vaiṣṇava traditions (Halbfass, 1991: 4).     

Policing Outlawed Desire: ‘Homocriminality’ in Beat Spaces in Australia

Drawing on interview data of gay men who have had their behavior in public spaces scrutinised by agents of the law for signs deviance, this article explores the historical characteristics of police animosity towards such conduct in Australia. This entails examining encounters between police and gay men who pursue desire in ‘beat’ (or ‘cottage’ to the use the UK term) spaces. Exploring why these outlaw gay male subjects are so abject and troubling to the law, the discussion documents how law’s desire to regulate gay men plays out in the masquerade of ‘plain-clothes’ agent provocateur operations where police entrap gay men by mimicking gay bodily appearances, gestures and mannerisms. This article also examines how police regulation of gay desire functions as a form of violence that delimits expressions of same sex desire in public spaces. A key theme that underpins the analyses in this paper is that the policing of desire in ‘beat’ spaces helps produce qualities of illicitness and dangerousness and that this, in turn, fuels the circuit of desire at play between gay men and agents of the law.

Interrupting the Myth of the <Emphasis Type="Italic">Partage</Emphasis>: Reflections on Sovereignty and Sacrifice in the work of Nancy,Agamben and Derrida

Agamben traces the bio-political essence of modern politics to the non-sacrificial killing of Homo Sacer in Roman law. Nancy, on the other hand, links the history of Western politics to the fundamental logic of sacrifice in Western metaphysics. He nevertheless contemplates the possibility that Western societies may finally have arrived at the threshold of a non-sacrificial existence. Derrida seeks to resist the sacrificial logic of Western metaphysics and politics, but nevertheless appears to accept it as an irreducible fact of human co-existence. Unlike Nancy, he envisages no actual or actualised beyond beyond the realm of sacrificial metaphysics and politics. He thus can be said to interrupt Nancy’s ‘myth’ of a non-sacrificial partage. This article compares these three philosophical stances in the hope of throwing more light on the role of sacrifice in the law and politics of our time. Professor of Law, Rand Afrikaans University. Conversations with Ann van Sevenant, Carol Clarkson, Louise du Toit, Peter Fitzpatrick, Costas Douzinas and Adam Thurschwell gave impetus to many of the themes developed in this article. Concomitant shortcomings and inaccuracies, as always, are mine.