Possible Nature of an Electron Dense Substance in the Thylakoid Lumen of Chloroplasts

The fixation of leaves of Tanacetum vulgare L. in glutaraldehyde makes it possible to isolate chloroplasts without sacrificing an electron dense substance in the thylakoid lumen. The extraction of lipids from the chloroplasts isolated from the leaves preliminarily fixed in glutaraldehyde has demonstrated that glycerolipids (galactolipids and phospholipids) are not manifested in TLC, whereas isoprenoid lipids (chlorophyll, carotenoids) are manifested. Presumably, isoprenoid lipids are not fixed with glutaraldehyde and are extracted from the thylakoid membrane. The ultrastructural control demonstrates that the electron dense substance from the thylakoid lumen is also extracted. It is possible that this substance is of isoprenoid nature.

Hrd1转基因小鼠的构建与验证

背景 内质网应激在糖尿病视网膜病变（DR）中发挥着重要作用.羟甲基戊二酰辅酶A还原酶降解蛋白1（Hrd1,又名Syvn1）作为一种内质网相关蛋白降解（ERAD）途径中的核心蛋白,能够减少内质网应激. 目的 构建Hrd1高表达小鼠,为今后研究Hrd1在DR中的作用提供动物模型. 方法 构建Hrd1系统表达重组质粒,酶切、测序后显微注射3 p1到685枚C57BL/6小鼠受精卵中,移植到代孕母鼠,得到Hrd1转基因小鼠.利用鼠尾PCR检测法鉴定F0代小鼠的基因型.得到Hrd1基因检测阳性鼠后,使其分别与野生型鼠杂交,得到F1代Hrd1转基因小鼠,取鼠尾组织,用PCR检测法再次鉴定F1代小鼠基因型. 结果 成功构建了Hrd1重组载体,重组质粒pRP.ExBi-EF1 a-Syvn1-IRES-eGFP的全序列测定结果显示,cDNA序列均正确.接受了Hrd1-pcDNA显微注射的685枚受精卵中成活598枚.将存活的受精卵移植到代孕母鼠后共产子鼠41只,获F0代子鼠8只,生理活动均良好.PCR电泳显示339 bp处阳性条带,该基因型可以传递到子代F1.结论 成功构建了Hrd1转基因小鼠.

Caspase-3 Mediates Apoptosis of Striatal Cells in GA Ⅰ Rat Model

In previous study,glutaric acid (GA) induced apoptosis of primary striatal neuron in vitro.In order to investigate the neurotoxic effects of GA on neonatal rat corpus striatum and the possible mechanism,34 male pups were randomly assigned to NS group,low dose GA (LGA,5 μmol GA/g body weight) group and high dose GA (HGA,10 μmol GA/g body weight) group.These pups were subcutaneously administered with three injections from postnatal day 3 to 22 at 7:30 am,15:00 pm and 22:30 pm and killed 12 h after the last injection.Microscopic pathology in corpus striatum was evalu-ated by HE staining.The apoptotic cells were identified by TUNEL staining.The transcript levels of caspase-3,8,9,Bax,Bcl-2 were detected by using real-time PCR and the protein levels of procaspase-3 and the active fraction were evaluated by Western blotting.In LGA and HGA groups,ventricle collapse,cortical atrophy by a macroscope and interstitial edema,vacuolations,widened perivascular space of bi-lateral striatum by a microscope were observed.TUNEL assay revealed that the apoptotic cells were in-creased in LGA and HGA groups.The transcript of caspase-3 was up-regulated to 2.5 fold,accompanied by the up-regulation of caspase-9,Bax and down-regulation of Bcl-2.The protein levels of procaspase-3 and the active fraction were up-regulated in LGA and HGA groups.The rat model for GA Ⅰ showed mitochondrial apoptotic pathway may be involved in the GA-induced striatal lesion.Further studies should be taken to investigate the underlying mechanisms.

Enzyme-Mediated Enantioselective Hydrolysis of Aliphatic Dicarboxylic Acid Diesters

The enzyme-mediated highly enantioselective hydrolysis of aliphatic dicarboxylic acid diesters has been developed. The racemic diesters were easily prepared by the coupling of racemic alcohols with dicarboxylic anhydrides followed by esterification or with dicarboxylic acids. In the cases of bis(1-phenylethyl) glutarate and bis(1-phenylethyl) adipate, the diesters which contained the dl- and meso-form diastereomers, were enantioselectively hydrolyzed by lipase from Candida antarctica (Novozym 435) in buffer at 30°C to afford the almost optically pure (R)-1-phenylethanol. On the other hand, the following chemical hydrolysis of the remaining (S, S)-diesters and (S)-monoesters gave the (S)-alcohol. Finally, both enantiomers were stoichiometrically obtained in about 100% isolated yield based on the racemic diesters. The enzymatic reaction was also applicable for the preparation of several optically active alcohols. In some cases, both the reactivities and enantioselectivities were quite different from those in the case of the corresponding simple acetates.

The effect of chemical structure of carboxylate molecules on hydroxyapatite nanoparticles.A structural and morphological study

Being the most abundant non-macromolecular organic component of bone,the role of citrate(Cit)in hydroxyapatite(HA)crystallization is of high relevance.In this work we have investigated the influence of hydroxycitrate(CitOH)and glutarate(Glr)on HA crystallization in terms of particle growth,composition,and morphology in comparison to Cit.CitOH and Glr have been selected for this work because they share the same backbone structure of Cit but bear different functional groups in the central region.Our data has revealed that CitOH strongly inhibits HA crystallization more efficiently than Cit.CitOH-HA nanoparticles are composed of platy,elongated particles similar to those of Cit-HA but they are ca.twice smaller and have a lower crystal order.On the other hand,Glr does not inhibit HA crystallization as Cit,but leads to the formation of OCP platelets that convert with maturation time to HA nanorods with larger aspect ratio than Cit-HA.In comparison to Cit-HA samples,Glr-HA nanoparticles have bigger dimensions,and higher structural order.Overall,our data reveal that the central carboxyl group of Cit is involved in the selective binding with HA crystal surface and in regulating HA crystal growth.The results of this work highlight new possibilities to control the formation of HA for designing advanced bioactive materials and give new insights on the role of the structure of Cit in regulating the HA morphology.

Recurrent rhabdomyolysis and glutaric aciduria type I: a case report and literature review

Background:Glutaric acidemia type I(GA-I)is a rare metabolic disorder caused by mutation of the glutaryl-CoA dehydrogenase(GCDH)gene.The occurrence of rhabdomyolysis with GA-I is extremely rare.Methods:We reported a child with recurrent rhabdomyolysis and undiagnosed glutaric acidemia type I(GA-I).And a literature review was performed.Results:A 4.5-year-old girl was admitted to our hospital due to recurrent rhabdomyolysis for 3 times within three years.At the third admission,she was diagnosed with GA-I by biochemical testing and mutation analysis.The girl was found to have a serine to leucine replacement mutation of the GCDH gene in exon 8 at position 764.Other three patients with rhabdomyolysis and GA-I were discovered by literature searching.Conclusion:This report highlights that patients with GA-I may have an increased risk of rhabdomyolysis.

Treatment of glutaric aciduria type I(GA-I)via intracerebroventricular delivery of GCDH

Glutaric aciduria type I(GA-I)is an autosomal recessive genetic disorder caused by a deficiency in glutaryl-CoA dehydrogenase(GCDH).Patients who do not receive proper treatment may die from acute encephalopathic crisis.Current treatments for GA-I include a low-lysine diet combined with oral supplementation of L-carnitine.A mouse model of Gcdh^(c.422_428del/c.422_428del)(Gcdh^(−/−))was generated in our laboratory using CRISPR/Cas9.Gcdh^(−/−)mice had significantly higher levels of glutaric acid(GA)in the plasma,liver,and brain than those in wild-type C57BL/6 mice.When given a high-protein diet(HPD)for two days,approximately 60%of Gcdh^(−/−)mice did not survive the metabolic stress.To evaluate whether GCDH gene replacement therapy could be used to provide sustained treatment for patients with GA-1,we prepared a recombinant adeno-associated virus(rAAV)carrying a human GCDH expression cassette and injected it into Gcdh^(−/−)neonates for a proof-of-concept(PoC)study.Our study demonstrated that delivering rAAV to the central nervous system(CNS),but not the peripheral system,significantly increased the survival rate under HPD exposure.Our study also demonstrated that rAAVPHP.eB mediated a higher efficiency than that of rAAV9 in increasing the survival rate.Surviving mice showed dose-dependent GCDH protein expression in the CNS and downregulation of GA levels.Our study demonstrated that AAV-based gene replacement therapy was effective for GA-I treatment and provided a feasible solution for this unmet medical need.

Condensation of α-and β-acetylnaphthalene with dimethyl β,β-dimethyl glutarate

α-and β-Acetylnaphthalenes condensed with dimethyl β,β-dimethyl glutarate in the presence of sodium hydride to give the corresponding half-esters,the E-isomers 2a and 2b being predominant. The structure and configuration of the half-esters were characterized by chemical and spectroscopic means.