蒙古黄鼠（Citellus dauricus）松果腺的超微结构观察

The distal part of pineal gland of the Mongolian ground squirrel was ultrastructurally studied. The gland was composed of low electron-dense parenchymal cells, among which glial cells, pigment cells, blood vessels and neural elements were occasionally interspersed. The pinealocytes contained numerous mitochondria, lysosomes, microtubules, microfilaments, Golgi apparatus and free ribosomes, as well as less prominent profiles of rough- and smooth-surfaced endoplasmic reticula and some cilia, centrioles, synaptic ribbons and few subsurface cisterns. Some pinealocytes were vacuolated. The content of the vacuoles released into the extracellular space by exocytosis could be observed. The gap junctions between pinealocytes were also observed. Of particular interest was that many mitochondria "fused together" and formed gap junction-like structure in about five percent pinealocytes. The pigment cell has a amorphous nucleus which contains many aggregated chromatin, its cell membrane has a few microvilli projecting into a central lumen, these features may indicated that this kind of cell differs either from the pinealocyte or astrocyte. There are axo-axonic synapses or axo-dendritic synapses between neuron processes or between neuron processes and pinealocytes.     

蒙古黄鼠(Citellus dauricus)松果腺的超微结构观察

蒙古黄鼠松果腺主要由低电子密度的松果腺细胞和少量的胶质细胞、含色素细胞、神经突起及血管等组成。松果腺细胞内含有大量的线粒体、溶酶体、微丝、高尔基器、游离核糖体及中等量的光面和粗面内质网。纤毛、中心粒、突触带和致密芯小泡很少。松果腺细胞之间及胶质细胞之间存在电突触。最新被观察到的是大约有5%松果腺细胞内的线粒体产生“融合”现象,形成类似电突触的结构。神经突起可形成轴—轴突触,轴—树突触,并与松果腺细胞形成突触。     

冬眠周期长短不同的蒙古黄鼠（Citellus dauricus）...

Adult Mongolian ground squirrels (Citellus dauricus) were kept at 5 degrees C in winter and divided into four experimental groups according to the bout length. The first group was not hibernating until decapitation. The bout length of the second group was between 4-10 days, the third group 11-17 days and the fourth group longer than 20 days. All pineals were sampled at the end of January. Morphometric analytical procedures were used to study the ultrastructure of the distal part of the pineal gland. The statistical results demonstrated that 1) the euthermic animals have larger cross areas of pinealocyte, longer and narrower Golgi apparatus and more number of saccules of each Golgi apparatus (P less than 0.01). But they also have smaller volume density of vaculoes, less lipid droplets and associated vesicles around Golgi apparatus (P less than 0.01). 2) the hibernating animals with variety of bout length had no significant differences in the number of mitochondria, lipid droplets, lysosomes, the size of Golgi apparatus and the cross areas of nucleus and cytoplasm (P greater than 0.05). However, the number and the cross areas of vacuoles were significantly increased with the bout length (P less than 0.01). This might suggest that the bout length was not related to the metabolic activity of pinealocytes in Citellus dauricus and vacuoles might play some important roles in maintenance of individual bout of hibernation in this species.     

Suppression of Lung Tumorigenesis by Leucine Zipper/EF Hand–Containing Transmembrane-1

Background

Leucine zipper/EF hand-containing transmembrane-1 (LETM1) encodes for the human homologue of yeast Mdm38p, which is a mitochondria-shaping protein of unclear function. However, a previous study demonstrated that LETM1 served as an anchor protein for complex formation between mitochondria and ribosome, and regulated mitochondrial biogenesis.

Methodology/Principal Findings

Therefore, we examine the possibility that LETM1 may function to regulate mitochondria and lung tumor growth. In this study, we addressed this question by studying in the effect of adenovirus-mediated LETM1 in the lung cancer cell and lung cancer model mice. To investigate the effects of adenovirus-LETM1 in vitro, we infected with adenovirus-LETM1 in A549 cells. Additionally, in vivo effects of LETM1 were evaluated on K-ras ^LA1 mice, human non-small cell lung cancer model mice, by delivering the LETM1 via aerosol through nose-only inhalation system. The effects of LETM1 on lung cancer growth and AMPK related signals were evaluated. Adenovirus-mediated overexpression of LETM1 could induce destruction of mitochondria of lung cancer cells through depleting ATP and AMPK activation. Furthermore, adenoviral-LETM1 also altered Akt signaling and inhibited the cell cycle while facilitating apoptosis. Theses results demonstrated that adenovirus-LETM1 suppressed lung cancer cell growth in vitro and in vivo.

Conclusions/Significance

Adenovirus-mediated LETM1 may provide a useful target for designing lung tumor prevention and treatment.     

Lamellar Gel (Lβ) Phases of Ternary Lipid Composition Containing Ceramide and Cholesterol

Lipid lateral segregation into specific domains in cellular membranes is associated with cell signaling and metabolic regulation. This phenomenon partially arises as a consequence of the very distinct bilayer-associated lipid physico-chemical properties that give rise to defined phase states at a given temperature. Until now lamellar gel (L_β) phases have been described in detail in single or two-lipid systems. Using x-ray scattering, differential scanning calorimetry, confocal fluorescence microscopy, and atomic force microscopy, we have characterized phases of ternary lipid compositions in the presence of saturated phospholipids, cholesterol, and palmitoyl ceramide mixtures. These phases stabilized by direct cholesterol-ceramide interaction can exist either with palmitoyl sphingomyelin or with dipalmitoyl phosphatidylcholine and present intermediate properties between raft-associated phospholipid-cholesterol liquid-ordered and phospholipid-ceramide L_β phases. The present data provide novel, to our knowledge, evidence of a chemically defined, multicomponent lipid system that could cooperate in building heterogeneous segregated platforms in cell membranes.     

“E-1059”对达乌尔黄鼠(Citellus dauricus)的毒性试验

“E-1059”是一种毒性大,而有内吸作用的杀虫剂。它的特点不仅杀虫力强,而且对哺乳动物的毒性也很大,因此把这种药品应用到鼠害防除方面,是大有可能的。特别是有些破坏草原的啮齿动物,不喜欢吃谷粒,如用内吸剂“E-1059”喷洒在草上来灭鼠,似乎很理想。但这首先要了解一下它对野鼠的毒性究竟怎样。     

Fabrication and In Vitro/In Vivo Performance of Mucoadhesive Electrospun Nanofiber Mats Containing α-Mangostin

This study aimed to fabricate mucoadhesive electrospun nanofiber mats containing α-mangostin for the maintenance of oral hygiene and reduction of the bacterial growth that causes dental caries. Synthesized thiolated chitosan (CS-SH) blended with polyvinyl alcohol (PVA) was selected as the mucoadhesive polymer. α-Mangostin was incorporated into the CS-SH/PVA solution and electrospun to obtain nanofiber mats. Scanning electron microscopy, differential scanning calorimetry, X-ray diffraction, and tensile strength testing were used to characterize the mats. The swelling degree and mucoadhesion were also determined. The nanofiber mats were further evaluated regarding their α-mangostin content, in vitro α-mangostin release, antibacterial activity, cytotoxicity, in vivo performance, and stability. The results indicated that the mats were in the nanometer range. The α-mangostin was well incorporated into the mats, with an amorphous form. The mats showed suitable tensile strength, swelling, and mucoadhesive properties. The loading capacity increased when the initial amount of α-mangostin was increased. Rapid release of α-mangostin from the mats was achieved. Additionally, a fast bacterial killing rate occurred at the lowest concentration of nanofiber mats when α-mangostin was added to the mats. The mats were less cytotoxic after use for 72 h. Moreover, in vivo testing indicated that the mats could reduce the number of oral bacteria, with a good mouth feel. The mats maintained the amount of α-mangostin for 6 months. The results suggest that α-mangostin-loaded mucoadhesive electrospun nanofiber mats may be a promising material for oral care and the prevention of dental caries.KEY WORDS: dental caries, mucoadhesive property, nanofibers, thiolated chitosan, α-mangostin     

Development of Acid-Resistant Alginate/Trimethyl Chitosan Nanoparticles Containing Cationic β-Cyclodextrin Polymers for Insulin Oral Delivery

In this study, the use of trimethylchitosan (TMC), by higher solubility in comparison with chitosan, in alginate/chitosan nanoparticles containing cationic β-cyclodextrin polymers (CPβCDs) has been studied, with the aim of increasing insulin uptake by nanoparticles. Firstly, TMCs were synthesized by iodomethane, and CPβCDs were synthesized within a one-step polycondensation reaction using choline chloride (CC) and epichlorohydrine (EP). Insulin–CβCDPs complex was prepared by mixing 1:1 portion of insulin and CPβCDs solutions. Then, nanoparticles prepared in a three-step procedure based on the iono-tropic pregelation method. Nanoparticles screened using experimental design and Placket Burman methodology to obtain minimum size and polydispercity index (pdI) and the highest entrapment efficiency (EE). CPβCDs and TMC solution concentration and pH and alginate and calcium chloride solution concentrations are found as the significant parameters on size, PdI, and EE. The nanoparticles with proper physicochemical properties were obtained; the size, PdI, and EE% of optimized nanoparticles were reported as 150.82 ± 21 nm, 0.362 ± 0.036, and 93.2% ± 4.1, respectively. The cumulative insulin release in intestinal condition achieved was 50.2% during 6 h. By SEM imaging, separate, spherical, and nonaggregated nanoparticles were found. In the cytotoxicity studies on Caco-2 cell culture, no significant cytotoxicity was observed in 5 h of incubation, but after 24 h of incubation, viability was decreased to 50% in 0.5 mμ of TMC concentration. Permeability studies across Caco-2 cells had been carried out, and permeability achieved in 240 min was 8.41 ± 0.39%, which shows noticeable increase in comparison with chitosan nanoparticles. Thus, according to the results, the optimized nanoparticles can be used as a new insulin oral delivery system.KEY WORDS: alginate, cationic β-cyclodextrin, insulin nanoparticle, oral delivery, trimethyl chitosan     

黄鼠(Citellus dauricus)基础代谢率、静止代谢率、化学热调节强度的季节性变化研究

本文测定了达乌尔黄鼠(Citellus dauricus)的耗氧量等指标,从代谢率与代谢调节方面探讨其能量代谢的季节性变化规律。结果表明,黄鼠的中性温度区在春、夏、秋各季有2—3℃的差别。此三个季节的基础代谢率(单位:毫升氧/克·小时,下同)分别为3.6182、2.6396、2.6005。春季显著高于夏季与秋季。在冬眠状态下室温为5℃时静止代谢率最低,为0.4436。在非冬眠各个季节,即0℃至下临界温度范围内静止代谢率与环境温度呈负相关并具线性回归。黄鼠的化学热调节强度较低,特别是在秋季和在冬眠状态下这种能力受到较深程度的抑制。黄鼠在非冬眠期的基础代谢率与体表面积有密切相关,冬眠期两者之间的相关不明显。     

冬眠周期长短不同的蒙古黄鼠(Citellus dauricus)松果腺细胞的形态研究

电镜观察和统计结果表明:1.不冬眠组蒙古黄鼠松果腺细胞相对冬眠组的来说具有较大截面积的细胞和核,较长且较窄的高尔基器,每个高尔基器所具有的囊数较多,但空泡的截面积小,脂肪粒少,透明小泡少;2.冬眠周期长短不同的黄鼠的松果腺细胞和核截面积大小无差异、高尔基器的大小也无差异,有差异的超微结构特征只有空泡的截面积大小,且它们三组之间的差异显著。冬眠周期越长,空泡的截面积越大。这提示蒙古黄鼠的冬眠周期长短可能与松果腺细胞的总代谢活力无关;空泡可能在维持冬眠周期长短中起重要作用。     

Nanosuspensions Containing Oridonin/HP-β-Cyclodextrin Inclusion Complexes for Oral Bioavailability Enhancement via Improved Dissolution and Permeability

Chemotherapy via oral route of anticancer drugs offers much convenience and compliance to patients. However, oral chemotherapy has been challenged by limited absorption due to poor drug solubility and intestinal efflux. In this study, we aimed to develop a nanosuspension formulation of oridonin (Odn) using its cyclodextrin inclusion complexes to enhance oral bioavailability. Nanosuspensions containing Odn/2 hydroxypropyl-β-cyclodextrin inclusion complexes (Odn-CICs) were prepared by a solvent evaporation followed by wet media milling technique. The nanosuspensions were characterized by scanning electron microscopy (SEM), differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy (FTIR), and dissolution. The resulting nanosuspensions were approximately 313.8 nm in particle size and presented a microcrystal morphology. Nanosuspensions loading Odn-CICs dramatically enhanced the dissolution of Odn. Further, the intestinal effective permeability of Odn was markedly enhanced in the presence of 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) and poloxamer. Bioavailability studies showed that nanosuspensions with Odn-CICs can significantly promote the oral absorption of Odn with a relative bioavailability of 213.99% (Odn suspensions as reference). Odn itself possesses a moderate permeability and marginal intestinal metabolism. Thus, the enhanced bioavailability for Odn-CIC nanosuspensions can be attributed to improved dissolution and permeability by interaction with absorptive epithelia and anti-drug efflux. Nanosuspensions prepared from inclusion complexes may be a promising approach for the oral delivery of anticancer agents.     

Discovery of Triterpenoids as Reversible Inhibitors of α/β-hydrolase Domain Containing 12 (ABHD12)

Background

α/β-hydrolase domain containing (ABHD)12 is a recently discovered serine hydrolase that acts in vivo as a lysophospholipase for lysophosphatidylserine. Dysfunctional ABHD12 has been linked to the rare neurodegenerative disorder called PHARC (polyneuropathy, hearing loss, ataxia, retinosis pigmentosa, cataract). In vitro, ABHD12 has been implicated in the metabolism of the endocannabinoid 2-arachidonoylglycerol (2-AG). Further studies on ABHD12 function are hampered as no selective inhibitor have been identified to date. In contrast to the situation with the other endocannabinoid hydrolases, ABHD12 has remained a challenging target for inhibitor development as no crystal structures are available to facilitate drug design.

Methodology/Principal Findings

Here we report the unexpected discovery that certain triterpene-based structures inhibit human ABHD12 hydrolase activity in a reversible manner, the best compounds showing submicromolar potency. Based on structure activity relationship (SAR) data collected for 68 natural and synthetic triterpenoid structures, a pharmacophore model has been constructed. A pentacyclic triterpene backbone with carboxyl group at position 17, small hydrophobic substituent at the position 4, hydrogen bond donor or acceptor at position 3 accompanied with four axial methyl substituents was found crucial for ABHD12 inhibitor activity. Although the triterpenoids typically may have multiple protein targets, we witnessed unprecedented selectivity for ABHD12 among the metabolic serine hydrolases, as activity-based protein profiling of mouse brain membrane proteome indicated that the representative ABHD12 inhibitors did not inhibit other serine hydrolases, nor did they target cannabinoid receptors.

Conclusions/Significance

We have identified reversibly-acting triterpene-based inhibitors that show remarkable selectivity for ABHD12 over other metabolic serine hydrolases. Based on SAR data, we have constructed the first pharmacophore model of ABHD12 inhibitors. This model should pave the way for further discovery of novel lead structures for ABHD12 selective inhibitors.     

Studies on γ-Radiolysis of Sulfur Containing Amino Acids

In connection with flavor deterioration accompanied by food irradiation, the effect of γ-irradiation on sulfoxide amino acids in air free aqueous system was investigated. The major radiolysis products from S-n-propyl-l-cysteine sulfoxide (PCSO) were alanine, cysteic acid, dipropyl disulfide, etc., and from S-allyl-l-cysteine sulfoxide (ACSO) were S-allyl-l-cysteine, cystine, cysteic acid, etc., which were isolated chromatographycally and identified by using IR and mass spectrometry. The sulfoxide in ACSO was more easily reduced to sulfide than that of in PCSO, and the bond of S-C (β-carbon in alanine moiety) in ACSO was difficult to cleave. These differences observed between PCSO and ACSO in the radiolysis products and their yields indicate that the radiolysis degradation is considerably influenced by the structure of alkyl group. From the experiments with N₂O or KBr addition during irradiation, principal roles of the active species in irradiated water in the degradation processes were partly elucidated.     

Synthesis of Oligonucleotides Containing 2′-Deoxyguanosine Adducts of Nitropyrenes

Two different approaches to synthesize oligonucleotides containing the 2 ′-deoxyguanosine adducts formed by nitropyrenes are described. A direct reaction of an unmodified oligonucleotide with an activated nitropyrene derivative is a convenient biomimetic approach for generating the major adducts in DNA. A total synthetic approach, by contrast, involves several synthetic steps, including Buchwald-Hartwig Pd-catalyzed coupling, but can be used for incorporating both the major and minor adducts in DNA in high yield.     

Nucleic Acid Analog Peptide Containing β-Aminoalanine Modified with Nucleobases

Abstract

Oligopeptides containing N^β-(thymin-1-ylacetyl) β-aminoalanine and N^β-(cytosin-1-ylacetyl) β-aminoalanine moieties synthesized on solid phase using standard boc-chemistry showed hybridization properties with single stranded DNA and RNA, and also with double stranded DNA at pH 7.0.     

Convenient Synthesis of Oligodeoxynucleotides Containing 2′-Deoxy-6-thioinosine

Abstract

A facile synthesis of oligodeoxynucleotides (ODN) containing 2′-deoxy-6-thioinosine (dI^6S) based on the convertible nucleoside O6-phenyl-2′-deoxyinosine is presented. After standard solid-phase DNA synthesis and removal of the cyanoethyl protecting groups with DBU treatment with aqueous sodium hydrogen sulfide introduces the sulfur functionality, deprotects the other nucleobases and cleaves the ODN from the solid support in a one-pot reaction. In addition, the extinction coefficient of 2′-deoxy-6-thioinosine is determined by enzymatic fragmentation of the resulting ODN in the presence of adenosine deaminase.     

α/β-Hydrolase Domain Containing Protein 15 (ABHD15) – an Adipogenic Protein Protecting from Apoptosis

Our knowledge about adipocyte metabolism and development is steadily growing, yet many players are still undefined. Here, we show that α/β-hydrolase domain containing protein 15 (Abhd15) is a direct and functional target gene of peroxisome proliferator-activated receptor gamma (PPARγ), the master regulator of adipogenesis. In line, Abhd15 is mainly expressed in brown and white adipose tissue and strongly upregulated during adipogenesis in various murine and human cell lines. Stable knockdown of Abhd15 in 3T3-L1 cells evokes a striking differentiation defect, as evidenced by low lipid accumulation and decreased expression of adipocyte marker genes. In preconfluent cells, knockdown of Abhd15 leads to impaired proliferation, which is caused by apoptosis, as we see an increased SubG1 peak, caspase 3/7 activity, and BAX protein expression as well as a reduction in anti-apoptotic BCL-2 protein. Furthermore, apoptosis-inducing amounts of palmitic acid evoke a massive increase of Abhd15 expression, proposing an apoptosis-protecting role for ABHD15. On the other hand, in mature adipocytes physiological (i.e. non-apoptotic) concentrations of palmitic acid down-regulate Abhd15 expression. Accordingly, we found that the expression of Abhd15 in adipose tissue is reduced in physiological situations with high free fatty acid levels, like high-fat diet, fasting, and aging as well as in genetically obese mice. Collectively, our results position ABHD15 as an essential component in the development of adipocytes as well as in apoptosis, thereby connecting two substantial factors in the regulation of adipocyte number and size. Together with its intricate regulation by free fatty acids, ABHD15 might be an intriguing new target in obesity and diabetes research.