有机磷酸酯引起的迟发性神经病的“强化”机理

某些酯酶抑制剂可以强化创伤和中毒引起的轴突病 ,这可以从临床症状和组织病理学上加以证实。有机磷酸酯诱导的迟发性神经病是研究轴突病强化机制的良好模型 ,有机磷酸酯诱导的迟发性神经病的强化可能与神经系统的修复机理有关。强化剂的靶标位点还未清楚 ,但不是神经病靶标酯酶 ,而可能与一种类似于神经病靶标酯酶的特殊酯酶有关     

氟丙胺磷对人成神经细胞瘤SK-N-SH细胞分化的毒性机制

目的研究诱发迟发性神经病的氟丙胺磷对人成神经细胞瘤SK-N-SH细胞分化的影响及其作用机理。方法MTT法测定细胞增殖,全反式维A酸诱导细胞分化,显微测量有机磷酸酯对细胞分化的影响,比色法测定NTE活力和Western印迹分析总的和磷酸化的神经纤丝重链蛋白以及肌动蛋白的表达。结果50μmol·L-1对氧磷和氟丙胺磷对细胞增殖没有影响;此浓度下对氧磷不抑制细胞的分化,而氟丙胺磷抑制细胞分化;氟丙胺磷抑制神经病靶标酯酶,而对氧磷对其不抑制;氟丙胺磷抑制总的和磷酸化的神经纤维丝重链蛋白的表达,而对氧磷对两种状态的神经纤维丝重链蛋白没有明显的作用。对氧磷和氟丙胺磷对肌动蛋白的表达都无影响。结论氟丙胺磷对人成神经细胞瘤细胞分化的抑制是通过对总的和磷酸化的神经纤维丝重链蛋白表达的抑制实现的,并伴随着神经病靶标酯酶的抑制。     

反向重复序列法构建神经病靶标酯酶RNA干涉表达质粒

目的构建基于通用真核表达载体的人神经病靶标酯酶双链RNA的稳定表达载体。方法在正义和反义引物两端分别加上EcoRⅠ和BamHⅠ的识别位点扩增得到神经病靶标酯酶活力域序列,构建含有目标基因的倒置重复序列的双链RNA表达载体pcDNA.NTE.dsRNA,酶切分析鉴定重组载体。脂质体法转染到Hela细胞中瞬时表达重组载体,酶活力测定其对细胞内神经病靶标酯酶活力的影响。结果成功构建了NTE双链RNA稳定表达载体pcDNA.NTE.dsRNA,表达该载体细胞内NTE活力明显下降至对照细胞的20%左右。结论采用反向重复序列法将靶标基因的编码序列正反向插入到通用真核细胞表达载体中,是构建哺乳细胞基因双链RNA稳定表达载体的有效方法。     

组蛋白脱乙酰酶抑制剂apicidin及其类似物的抗原虫与抗肿瘤作用研究进展

Apicidin是镰刀菌代谢产物,属环四肽类分子,能结合组蛋白脱乙酰酶(HDAC)使组蛋白过度乙酰化,具有广谱抗顶复亚门原虫和广谱抗肿瘤作用,可抑制肿瘤细胞转移、增殖和促使肿瘤细胞凋亡、分化.对apicidin类似物的生物学活性研究表明,apicidin的癸酸侧链、大环结构和色氨酸侧链是药效基团元件,使其能竞争性结合HDAC,发挥其生物学效应.     

青霉素单克隆抗体研究——青霉素类抗生素交叉过敏反应及其抗原决定簇

利用酶联免疫吸附试验研究4个青霉素单克隆抗体和5种不同侧链青霉素细胞色素C结合物作用,探讨青霉素交叉过敏反应及其抗原决定簇.实验证明青霉素过敏反应抗原决定簇——青霉噻唑基团,有二个结合位点,一是可被D8单克隆抗体所识别的青霉素侧链,一是可被D44、D7等单抗识别的青霉素β-内酰胺环破裂后与载体上氨基结合形成新位点.我们认为新位点是青霉素类抗生素间的交叉过敏反应的重要位点.头孢噻吩不具有四氢味喃环而其与细胞色素C结合物与单抗D8、D44、D7亲和力仅弱于BPO_4—Cy,又从竞争性抑制酶联免疫吸附试验结果亦表明四氢呋喃环不是一个结合位点,头孢噻吩和青霉素间交叉反应由于有共同新结合位点是重要因素,所以我们推测青霉素及头孢霉素间可能存在着交叉过敏反应.     

抗生素的细菌耐药性：酶降解和修饰

抗生素耐药性通过三种机制而起作用:阻止药物与靶标相互作用;从细胞中排出抗生素;对抗生素的直接破坏和修饰。本文主要讨论抗生素的直接破坏和修饰而失活,这包括:水解、基团转移和氧化还原机制。而水解对于临床非常重要,特别是β-内酰胺类抗生素应用以后。而基团转移有多种途径,包括乙酰基转移修饰、磷酸化、糖基化、核苷酸化、核糖基化和巯基转移。酶对抗生素修饰的唯一特点是,这些机制单独起作用降低了药物在局部环境中的浓度,因而,药物研发者和临床医生面临的挑战是针对这种机制找到抗感染治疗的新方法。本文将概括目前有关抗生素耐药性的一些研究成果,并讨论耐药性酶的分子机制、三维结构及进化,从而克服抗生素的耐药性。     

乙酰肝素酶促进肿瘤转移分子机制的最新研究进展

乙酰肝素酶是目前发现的哺乳动物细胞中唯一能切割细胞外基质中硫酸肝素蛋白多糖侧链的内源性糖苷酶,参与许多病理及生理的过程。本文对乙酰肝素酶与肿瘤转移的高度相关性以及该酶参与肿瘤转移的分子机制等方面进行了综述,乙酰肝素酶会通过促进肿瘤血管生成、影响细胞分化以及促进肿瘤细胞黏附作用和侵袭能力等途径促进肿瘤的播散,因此可以将乙酰肝素酶作为抗肿瘤药物的靶点。本文对用筛选乙酰肝素酶抑制剂的方法、寻找抗肿瘤新药的最新进展做了概述,旨在为肿瘤转移机制研究,寻找治疗方法提供一些线索。     

一种新型流感病毒神经氨酸酶抑制剂的设计、合成及活性研究

本研究以H5N1亚型流感病毒神经氨酸酶(NA)活性位点旁的150-空穴为靶标,采用半柔性分子对接计算机模拟技术,设计并合成了一种绿原酸的结构类似物——4-(咖啡酰基)氨基丁酸,计算机模拟结果显示该化合物能够插入到N1150-空穴中并和Arg156侧链以氢键的方式结合,与N1的最佳结合自由能为-7.70 kcal.mol-1,与奥司他韦相当。同时,利用以H5N1假病毒体系为基础建立的NA抑制剂筛选模型,测定了奥司他韦、绿原酸和4-(咖啡酰基)氨基丁酸对NA的抑制作用,发现与绿原酸相比,4-(咖啡酰基)氨基丁酸显著增强了对N1型神经氨酸酶的抑制作用,但与奥司他韦仍有一定的差距。本实验初步探索了150-空穴作为新型神经氨酸酶抑制剂靶标的可能性,为开发新型神经氨酸酶抑制剂提供了新的思路。     

荧光标记多糖的技术及其应用

多糖是生命科学中不可缺少的生物大分子,由于其本身缺少易于检测的发光基团,一直困扰着人们对多糖进一步的研究。荧光物质具有荧光发光基团,通过化学合成的方法将其结合到糖链分子上,从而使多糖带有荧光基团,以便于我们用常用的检测手段进行检测,从而进一步了解多糖发挥的作用,并对其作用机制进行研究。     

灵芝酸抗癌机制的分子模拟研究

目的分析灵芝酸与肿瘤靶酶的相互作用模式,研究灵芝酸的抗肿瘤机制。方法根据灵芝酸结构寻找可能的肿瘤靶点,并基于靶点结构,利用分子对接,探讨灵芝酸与靶酶的相互作用模式以及作用强度。结果与结论预测了灵芝酸与肿瘤靶标的结合方式,有助于理解灵芝酸的抗肿瘤机制。     

The homeostasis of phosphatidylcholine and lysophosphatidylcholine was not disrupted during tri-o-cresyl phosphate-induced delayed neurotoxicity in hens

Little is known regarding early biochemical events in organophosphate-induced delayed neurotoxicity (OPIDN) except for the essential inhibition of neuropathy target esterase (NTE). We hypothesized that the homeostasis of lysophosphatidylcholine (LPC) and/or phosphatidylcholine (PC) in nervous tissues might be disrupted after exposure to the organophosphates (OP) which participates in the progression of OPIDN because new clues to possible mechanisms of OPIDN have recently been discovered that NTE acts as lysophospholipase (LysoPLA) in mice and phospholipase B (PLB) in cultured mammalian cells. To bioassay for such phospholipids, we induced OPIDN in hens using tri-o-cresyl phosphate (TOCP) as an inducer with phenylmethylsulfonyl fluoride (PMSF) as a negative control; and the effects on the activities of NTE, LysoPLA and PLB, the levels of PC, LPC, and glycerophosphocholine (GPC), and the aging of NTE enzyme in the brain, spinal cord, and sciatic nerves were examined. The results demonstrated that the activities of NTE, NTE-LysoPLA, LysoPLA, NTE-PLB and PLB were significantly inhibited in both TOCP- and PMSF-treated hens. The inhibition of NTE and NTE-LysoPLA or NTE-PLB showed a high correlation coefficient in the nervous tissues. Moreover, the NTE inhibited by TOCP was of the aged type, while nearly all of the NTE inhibited by PMSF was of the unaged type. No significant change in PC or LPC levels was observed, while the GPC level was significantly decreased. However, there is no relationship found between the GPC level and the delayed symptoms or aging of NTE. All results suggested that LPC and/or PC homeostasis disruption may not be a mechanism for OPIDN because the PC and LPC homeostasis was not disrupted after exposure to the neuropathic OP, although NTE, LysoPLA, and PLB were significantly inhibited and the GPC level was remarkably decreased.     

USING NEUROBLASTOMA CELL LINES TO ADDRESS DIFFERENTIAL SPECIFICITY TO ORGANOPHOSPHATES

1. Organophosphates can cause acute toxicity, which follows inhibition of ncetylcholinesterase (AChE), or delayed neuropathy, which follows inhibition of neuropathy target esterase (NTE). 2. Human neuroblastoma SH-SY5Y cells contain AChE and NTE. 3. Organophosphates actively able to inhibit AChE in animal models inhibited AChE in neuroblastoma cells. 4. Inhibition of NTE in neuroblastoma cells could identify active organophosphates capable of causing delayed neuropathy in animal models and distinguish these organophosphates from those that do not cause delayed neuropathg in animal models.     

Clinical expression of organophosphate-induced delayed polyneuropathy in rats.

Single doses of certain organophosphates (OP), such as dibutyl-2,2-dichlorovinyl phosphate (DBDCVP) cause organophosphate-induced delayed polyneuropathy (OPIDP) in hens. Clinical effects correlate with inhibition of neuropathy target esterase (NTE) which is considered the target for this toxicity. Pre-treatment with non-neuropathic NTE inhibitors, such as phenylmethanesulfonyl fluoride (PMSF), protects from OPIDP. However, when given after OPs, these compounds promote OPIDP. Chicks are relatively resistant to OPIDP despite high NTE inhibition. It has also always been reported that rats represent a species which is resistant to OPIDP and that they might develop morphological but not clinical signs of OPIDP. We report here that clinical OPIDP can be produced in 3.5- and 6-month-old rats by DBDCVP (5 mg/kg s.c.) and that it correlates with high (> 90%) NTE inhibition. When PMSF (120 mg/kg s.c. x 2) was given after DBDCVP, OPIDP was promoted. Pretreatment with PMSF protected from OPIDP. We conclude that resistance to OPIDP in the rat is age-related, as it is in the hen.     

Phenylmethanesulfonyl fluoride elicits and intensifies the clinical expression of neuropathic insults

It has been recently reported that phenyl-methanesulfonyl fluoride (PMSF) when given to hens after a neuropathic organophosphate (OP) promotes organophosphate-induced delayed polyneuropathy (OPIDP). Chicks are resistant to OPIDP despite high inhibition/aging of neuropathy target esterase (NTE), the putative target of OPIDP initiation. However, when PMSF (300 mg/kg s.c.) is given to chicks after di-butyl 2,2-dichlorovinyl phosphate (DBDCVP, 1 or 5 mg/kg s.c.), OPIDP is promoted. Inhibition/aging of at least 30% of NTE was thought to be an essential prerequisite for promotion to be elicited in adult hens. However, we observed in hens that when NTE is maximally affected (>90%) by phenyl N-methyl N-benzyl carbamate (40 mg/kg i.V.), a non-ageable inhibitor of NTE, and then PMSF is given (120 mg/kg/day s.c. × 3 days) clinical signs of neuropathy become evident. Methamidophos (50 mg/kg p. o. to hens), which produces in vivo a reactivatable form of inhibited NTE, was shown either to protect from or promote OPIDP caused by DBDCVP (0.45 mg/kg s. c), depending on the sequence of dosing. Because very high doses of methamidophos cause OPIDP, we considered this effect to be a “self-promoted” OPIDP. We concluded that NTE inhibitors might have different intrinsic activities for producing OPIDP once NTE is affected. Aging might differentiate highly neuropathic OPs, like DBDCVP, from less neuropathic OPs, like methamidophos, or from the least neuropathic carbamates, which require promotion in order for neuropathy to be expressed. Retrograde axonal transport in motor fibers was measured as the accumulation of¹²⁵ I-tetanus toxin in spinal cord after injection in the gastrocnemius muscle of chicks treated either with DBDCVP (5 mg/kg s.c.) or with DBDCVP followed by PMSF (300 mg/kg s.c). Retrograde axonal transport was reduced in both groups (to about 50%, 10 days after dosing) and returned to normal 27 days after dosing. However, DBDCVP-treated chicks had a mild neuropathy which recovered relatively quickly, whereas chicks to which PMSF was also given had more severe signs which did not recover by day 27. We concluded that promotion affects a site other than NTE and that it acts at a point downstream from initiation. PMSF was also shown to promote 2,5-hexanedione (2,5-HD) neuropathy. 2,5-HD was given to hens at doses (200 mg/kg/day i.p. × 8 days) which caused mild and reversible neuropathy. When PMSF (120 mg/kg/day × 2 days at the end of 2,5-HD treatment) was given, more severe and irreversible signs of neuropathy were observed. We conclude that promotion might be a common feature in neuropathies of different origin. Part of this work was presented at the 30th Annual Meeting of the Society of Toxicology held in Dallas, TX, USA, February – March 1991     

Organophosphate polyneuropathy and neuropathy target esterase: Studies with methamidophos and its resolved optical isomers

 Methamidophos (O, S-dimethyl phospho rothioamidate) causes polyneuropathy in man and hens. However, experiments in the hen show that lower doses of methamidophos either protect from or promote the neuropathy caused by certain organophosphates. The initiation of neuropathy as well as protection from neuropathy are thought to be related to neuropathy target esterase (NTE), whereas promotion is likely to be due to interactions with another unknown target. Methamidophos is a racemate and we report studies with its resolved optical isomers, aimed at elucidating which isomer is responsible for the described effects. The time-course of acetylcholinesterase (AChE) and NTE activity in nervous tissues of hens after inhibition by single doses of either isomer showed that after D-(+) methamidophos (25 mg/kg PO) peak inhibition of both enzymes was achieved within 24 h (80–90%). However, after L-(−) methamidophos (15 mg/kg PO), peak inhibition (80–90%) was obtained within 24 h for AChE, whereas similar NTE inhibition (120 mg/kg PO) was observed only 4 days after dosing. The minimal neuropathic doses of D-(+) and L-(−) methamidophos were 60 and 120 mg/kg PO, respectively, and correlated with >80% NTE inhibition in nervous tissues. OPIDP initiation by either isomer was slightly promoted by phenylmethanesulfonyl fluoride (120 mg/kg SC). D-(+) Methamidophos (25 mg/kg PO) partially protected from dibutyl dichlorovinylphosphate (DBDCVP) neuropathy (up to 0.8 mg/kg SC). This effect correlated with about 70% NTE inhibition. L-(−) Methamidophos (15 or 60 mg/kg PO) did not protect from DBDCVP neuropathy (0.2–0.8 mg/kg SC). D-(+) and L-(−) methamidophos (25 mg/kg PO) promoted DBDCVP neuropathy (0.4 mg/kg SC), and D-(+) methamidophos (24 mg/kg PO) also promoted DFP neuropathy (0.3 mg/kg SC). These effects were unrelated to the degree of NTE inhibition they caused: about 70% by D-(+) methamidophos and extrapolated to about 10–15% by L-(−) methamidophos. We conclude that when racemic methamidophos is given to hens, initiation and protection from OPIDP is due to the interaction of D-(+) methamidophos with NTE. Promotion of OPIDP is due to both isomers as the result of their interaction with unknown site(s). It is possible that the neuropathy due to racemic methamidophos or isomers is a self promoted neuropathy because the promoting doses of both isomers are much lower than the neuropathic ones, and because the neuropathy they initiate is only slightly promoted by phenylmethanesulfonyl fluoride. Received: 5 July 1994/Accepted: 17 October 1994     

Sensitivity and selectivity of compounds interacting with neuropathy target esterase. Further structure-activity studies

Assay of neuropathy target esterase (NTE) which accounts for about 70% of paraoxon-resistant phenyl valerate (PV) esterase activity of hen brain depends on the fact that it is selectively inhibited by mipafox. A previous study of structure/activity relationships (Biochem. Pharmac. 24, 797, 1975) has been extended. Among 14 potential substrates NTE hydrolysed phenyl phenoxyacetate and phenyl thiophenoxyacetate faster (1.5-1.7X) than PV, but selectivity of these substrates for NTE among the paraoxon-resistant esterases was only 35-52%. Seventy-seven other potential inhibitors (organophosphates, phosphonates, phosphoramidates, phosphinates and carbamates) were examined to determine I50NTE and effects on both NTE and "non-NTE" at 3-4 x I50NTE (I 85-95) and, where possible, at 6-20 X I50NTE. Hydrophophic inhibitors with small/flexible leaving groups were generally very inhibitory: several 2,2-dichlorovinyl phosphates and fluorides were active at low nanomolar concentrations. In the dichlorovinyl phosphate series increasing dialkyl chain length beyond n-pentyl decreased inhibitory power, presumably due to steric hindrance since the methyl/n-decyl ester was 15X more active than di-n-decyl. Chloro-substitution of both ortho-positions of a phenyl leaving group for benzylcarbamates reduced inhibitory power more than 20X but had little effect in a phenyl leaving group of methyl phenylphosphonates where the acyl-leaving group bond is longer and less subject to steric hindrance. N-phenylbenzohydroxamyl benzylcarbamate is 10X more potent than any previously described carbamate against NTE. Among stereo-isomers differences of activity ranged from less than 2- to 15-fold. Only diphenylphosphinyl fluoride appeared to be virtually specific for NTE: at 0.5-1 microM it inhibited ca.92% of NTE and 10-13% of "non-NTE" which is similar to the specificity found for 2,6-dichlorophenyl methyl phenylphosphonate which has been claimed to be specific. Diphenylphosphinyl fluoride has an advantage in that it is easily synthesized and should be protective rather than neuropathic, but it is not stable in store. We cannot repeat experiments purporting to show a substantial proportion of a second isozyme of NTE. However, according to first-order kinetics, concentrations of inhibitor greater than 6 X I50 should inhibit NTE greater than 98% and for 19 out of 26 compounds a residue greater than 3% (limit of precision) was found under these conditions: in nearly every case the quantity was 3-5%. This quantity may not be "true NTE" but it cannot be the target for organophosphate-induced delayed neuropathy since it is resistant to various neuropathic and protective compounds.(ABSTRACT TRUNCATED AT 400 WORDS)     

In vitro modifications of rat NTE and other esterases by chemicals which induce delayed neurotoxicity in vivo

A rat in vitro model has been developed which permits direct study of the biochemical mechanisms involved in delayed neurotoxicity induced by any chemical compound, not only organophosphates. Using rat brain homogenate, a parallel study on the activity of neurotoxic esterase (NTE) and total esterases (TE) compared the action of metamidophos, which is known to induce delayed neurotoxicity, and the synthetic fatty acid anilides, oleylanilide and linoleylanilide. Inhibition in the activity of NTE and TE, unrelated to the concentration and the incubation time assayed, was caused by metamidophos, while the anilides showed a 2-phase concentration-time dependent behaviour. This confirmed the results we previously obtained in vivo. In both cases the appearance of delayed neuropathy was related to modification of NTE activity. We concluded that phosphorylation of the enzyme may not be the only biochemical requirement for the development of delayed neurotoxicity syndromes in which modification of NTE is produced.     

In vivo and in vitro regional differential sensitivity of neuropathy target esterase to Di-n-butyl-2,2-dichlorovinyl phosphate

Organophosphate-induced delayed polyneuropathy (OPIDP) is initiated by inhibition/aging of more than 70–75% of neuropathy target esterase (NTE). Di-n-butyl-2,2-dichlorovinyl phosphate (DBDCVP) (1 mg/kg s.c.) inhibited 96%, 86% and 83% of NTE in brain, spinal cord and peripheral nerve, respectively, and induced a typical central peripheral distal axonopathy in hens. A lower dose (0.45 mg/kg s.c.) caused 90%, 83% and 54% NTE inhibition in the same organs; by contrast, hens developed a spastic ataxia with axonal degeneration in spinal cord but not in peripheral nerve. With a dose of 0.2 mg/kg s.c., a suprathreshold inhibition of NTE was produced in brain (78%) but not in spinal cord (56%) and peripheral nerve (33%) and no morphological or clinical signs of neuropathy developed in hens. With doses up to 4.0 mg/kg s.c., acetylcholinesterase (AChE) inhibition was similar throughout the nervous system. In vitro time-course inhibition studies showed a different sensitivity to DBDCVP of NTE from peripheral nerve (k_a = 5.4 × 10⁶) relative to that from spinal cord (k_a = 13.9 × 10⁶) or brain (k_a = 20.6 × 10⁶). In vitro I₅₀s of DBDCVP for AChE were similar in brain, spinal cord and peripheral nerve (11–17 nM). These data support the hypothesis that the critical target for initiation of OPIDP is located in the nerve fiber, possibly in the axon and also suggest that peripheral nerve NTE has a different sensitivity to DBDCVP than the brain enzyme. Moreover, they confirm data showing that the degree of NTE inhibition in brain after dosing with organophosphates may not be a good monitor for the enzyme in parts of the nervous system where axonal degeneration actually develops. Therefore, direct assay of peripheral nerve NTE yields data which closely correlate with degree of axonal degeneration.Part of this work was presented at the 26th Annual Meeting of the Society of Toxicology, held in Washington DC, USA, February 24–27, 1987 and at the International Meeting on Esterases, Hydrolysing Organophosphorus Compounds, held in Dubrovnik, Yugoslavia, April 24–27, 1988     

Central-peripheral delayed neuropathy caused by diisopropyl phosphorofluoridate (DFP): segregation of peripheral nerve and spinal cord effects using biochemical, clinical, and morphological criteria

Systemic injection of diisopropyl phosphorofluoridate (DFP; 1 mg/kg, sc) causes delayed neuropathy in hens. This effect is associated with a high level of organophosphorylation of neuropathy target esterase (NTE) followed by an intramolecular rearrangement called "aging." Phenylmethanesulfonyl fluoride (PMSF) also attacks the active center of NTE but "aging" cannot occur. This compound does not cause neuropathy and protects against a subsequent challenge systemic dose of DFP. Intraarterial injection of DFP (0.185 mg/kg) into only one leg of hens caused a high NTE inhibition (greater than 80%) in the sciatic nerve of the injected leg, but not in other parts of the nervous system (37% average). A unilateral neuropathy with typical histopathological lesions developed in the injected leg. PMSF (0.55 mg/kg) injected into each sciatic artery caused 47% inhibition of sciatic nerve NTE but only 17-22% inhibition of NTE elsewhere; it did not produce clinical or histopathological lesions. When these hens were challenged with DFP (1 mg/kg, sc), high inhibition of residual-free NTE (greater than 85%) occurred throughout the nervous system and clinical signs of a syndrome different from the classical delayed neuropathy developed: this spinal cord type of ataxia was associated with histopathological lesions in the spinal cord but not in peripheral nerve. PMSF (1 mg/kg) injected into only one sciatic artery caused selective protective inhibition of sciatic nerve NTE of that leg. After systemic challenge by DFP, clinical effects expressed were a combination of spinal cord ataxia plus unilateral peripheral neuropathy. The challenge dose of DFP (1 mg/kg, sc) was insufficient to produce clear histopathological lesions in unprotected peripheral nerves although spinal lesions were found in these hens. Thus clinical evaluation of the peripheral nervous system by means of walking tests and a simple test of "leg retraction" reflexes was more sensitive and specific in diagnosis of peripheral neuropathy than was the histopathology.     

Announcement

Chymotrypsin and neurotoxic esterase (NTE) have some similarities. After inhibition of concentrated (80–800 μM) chymotrypsin by aryl saligenin cyclic phosphates it is known that aging occurs and some phenolic material becomes attached to protein. This binding has now been shown to be a manifestation of non-specific reaction with any available electrophile such as Tris, reduced glutathione (GSH), or protein. The reaction is therefore not a model for the 100% efficient transfer of alkyl groups to protein which occurs during aging of NTE inhibited by dialkyl phosphates.