Correction: Blood brain barrier permeable gold nanocluster for targeted brain imaging and therapy: an in vitro and in vivo study.

Correction for 'Blood brain barrier permeable gold nanocluster for targeted brain imaging and therapy: an in vitro and in vivo study' by L. V. Nair et al., J. Mater. Chem. B, 2017, 5, 8314-8321, https://doi.org/10.1039/C7TB02247F.

Additional muscle fibers of brachioradialis with anomalous high origin and entrapment of radial nerve in an osseomuscular canal.

Normally, brachioradialis originates from the upper part of the lateral supracondylar ridge of the humerus. Variations in its origin are very rare. We observed the presence of an additional set of fleshy muscle fibers in the lateral part of the anterior compartment of the arm in addition to the other normal muscles. This unusual case was observed at the Mookambika Institute of Medical Sciences, Kulashekara, Tamil Nadu, India, during routine dissection of the upper limb. The anomalous fleshy fibers were attached proximally to the middle part of the shaft of the humerus, close to the insertion of deltoid. Some of its fibers continued further up to the acromian process of scapula. These fibers passed downwards along with deltoid and joined the fibers from the humerus before getting merged with the brachioradialis distally. These additional muscle fibers were compared with the brachioradialis accessorius and the uniqueness, functional significance and the clinical relevance werediscussed (Fig. 2, Ref. 6). Full Text in PDF www.elis.sk.

Blood brain barrier permeable gold nanocluster for targeted brain imaging and therapy: an in vitro and in vivo study.

Blood brain barrier (BBB) is a dynamic interface, comprising polarized endothelial cells, that separates the brain from the circulatory system. The highly protective nature of this tight junction impairs diagnosis and treatment of brain disorders. In this study, we designed a sub atomic size, near infrared emitting, dual function glutathione gold cluster with high fluorescence yield to facilitate permeability of BBB, for imaging applications and drug delivery. The gold cluster was then modified with Levodopa (l-dopa), to utilize the large amino acid transporter 1 (LAT1) pathways to enhance brain entry. Uptake and permeability of the nanoprobes were demonstrated using an established model of BBB, comprising brain endothelial cells (bEnd.3). The uptake and the clearance of l-dopa modified cluster was faster than the glutathione cluster. l-Dopa modified cluster supports the slow and sustained delivery of a model drug, pilocarpine, to the brain. Results of in vivo imaging and drug release in normal mice hold promise for considering the probe for early diagnosis of brain diseases, when the barrier is not disrupted, and for subsequent drug treatment.

Sequence and arrangement of two genes of the butyrate-synthesis pathway of Clostridium acetobutylicum ATCC 824.

The genes encoding both Clostridium acetobutylicum ATCC 824 butyrate synthesis pathway enzymes, phosphotransbutyrylase (ptb) and butyrate kinase (buk), were sequenced. The genes are immediately adjacent on the chromosome, with ptb preceding buk. A single transcription start point (tsp) was identified 57 bp upstream from the ptb start codon by primer extension analysis. The ptb and buk genes appear to form an operon. A putative Rho-independent terminator structure was identified 26 bp downstream from buk.

Synthesis and antimetastatic properties of stereoisomeric tricyclic bis(dioxopiperazines) in the Lewis lung carcinoma model.

Synthesis of trans- and cis-tetrahydrodipyrazino[1,2-a:1',2'-d] pyrazine-1,3,7,9(2H,4H,8H,10H)-tetrone analogues 10 and 11 belonging to the bis(dioxopiperazine) class of antitumor agents and their bis(morpholinomethyl) derivatives 12 and 13 are described with use of 2,5-dimethylpyrazine as the starting material. Synthetic studies utilizing 3,6-disubstituted 2,5-dioxopiperazine precursors are included. Evaluation of 10-13 in the Lewis Lung carcinoma model indicated the bis(morpholinomethyl) analogue cis-13 to be antimetastatic, whereas the trans isomer 12 was toxic at a similar dose effecting a decrease in the life span of treated mice. The parent bis(dioxopiperazines) 10 and 11 were ineffective as antitumor or antimetastatic drugs.

Inhibition of smooth muscle cell proliferation in vitro by leflunomide, a new immunosuppressant, is antagonized by uridine.

Chronic rejection in the form of graft vascular disease (GVD) continues to plague clinical transplantation of vascularized organs. The histopathology of this lesion is characterized by neointimal hyperplasia, smooth muscle cell proliferation, and obliterative arteriopathy. Due to the lack of effective medical therapy for preventing or reversing these chronic vascular changes, retransplantation remains the final resort in treatment. Some of the newer immunosuppressive agents, including the new isoxazole derivative leflunomide (LFM), have shown efficacy in preventing chronic rejection in animal models of transplantation. Although its mechanism of action remains incompletely elucidated, previous work using lymphocytes in vitro suggests that the drug might act as a tyrosine kinase inhibitor, an inhibitor of de novo pyrimidine biosynthesis, or both. In order to elucidate whether the efficacy of LFM in vivo is attributable not only to anti-proliferative effects on the recipient immune system but also to direct effects on mesenchymal cells in the donor organ, we examined the effects of LFM on a transformed 9E11G murine smooth muscle cell (M-SMC) line in vitro. We demonstrate here that the active metabolite of LFM, A77 1726, dose-dependently inhibits the constitutive and growth-factor stimulated proliferation of M-SMC in vitro. Furthermore, the anti-proliferative effect of the drug can be reversed by the addition of uridine to the culture medium. These results suggest that inhibition of uridine biosythesis appears to be a mechanism by which LFM exerts anti-proliferative effects on both lymphocytes and smooth muscle cells, and this dual action may be responsible for its efficacy in preventing GVD in vivo.

Inhibition of smooth muscle cell proliferation in vitro by leflunomide, a new immunosuppressant, is antagonized by uridine.

Chronic rejection in the form of graft vascular disease (GVD) continues to plague clinical transplantation of vascularized organs. The histopathology of this lesion is characterized by neointimal hyperplasia, smooth muscle cell proliferation, and obliterative arteriopathy. Due to the lack of effective medical therapy for preventing or reversing these chronic vascular changes, retransplantation remains the final resort in treatment. Some of the newer immunosuppressive agents, including the new isoxazole derivative leflunomide (LFM), have shown efficacy in preventing chronic rejection in animal models of transplantation. Although its mechanism of action remains incompletely elucidated, previous work using lymphocytes in vitro suggests that the drug might act as a tyrosine kinase inhibitor, an inhibitor of de novo pyrimidine biosynthesis, or both. In order to elucidate whether the efficacy of LFM in vivo is attributable not only to anti-proliferative effects on the recipient immune system but also to direct effects on mesenchymal cells in the donor organ, we examined the effects of LFM on a transformed 9E11G murine smooth muscle cell (M-SMC) line in vitro. We demonstrate here that the active metabolite of LFM, A77 1726, dose-dependently inhibits the constitutive and growth-factor stimulated proliferation of M-SMC in vitro. Furthermore, the anti-proliferative effect of the drug can be reversed by the addition of uridine to the culture medium. These results suggest that inhibition of uridine biosynthesis appears to be a mechanism by which LFM exerts anti-proliferative effects on both lymphocytes and smooth muscle cells, and this dual action may be responsible for its efficacy in preventing GVD in vivo.

Obliterative bronchiolitis after lung and heart-lung transplantation.

Obliterative bronchiolitis (OB) has emerged as the main cause of morbidity and mortality in the long-term follow-up after lung and heart-lung transplantation. The pathogenesis of OB is multifactorial, with acute rejection and cytomegalovirus infection being the main risk factors for the development of OB. The final common pathway of all inciting events seems to be an alloimmune injury, with subsequent release of immunologic mediators and production of growth factors leading to luminal obliteration and fibrous scarring of the small airways. Analyzing the 14 years of experience in 163 patients at Stanford University, we found a current incidence of bronchiolitis obliterans syndrome or histologically proven OB within the first 3 years after lung and heart-lung transplantation of 36.3%, with an overall prevalence of 58.1% after heart-lung and 51.4% after lung transplantation. Both pulmonary function indices (forced expiratory flow between 25% and 75% of forced vital capacity and forced expiratory volume in 1 second) and transbronchial biopsies have proven helpful in diagnosing bronchiolitis obliterans syndrome or OB at an early stage. Early diagnosis of OB and improved management have achieved survival rates in patients with OB after 1, 3, 5, and 10 years of 83%, 66%, 46%, and 22%, compared with 86%, 83%, 67%, and 67% in patients without OB. Recently, different experimental models have been developed to investigate the cellular and molecular events leading to OB and to evaluate new treatment strategies for this complication, which currently limits the long-term success of heart-lung and lung transplantation.

Adaptation of solid phase extraction to an automated column switching method for online sample cleanup as the basis of a facile and sensitive high-performance liquid chromatographic assay for paclitaxel in human plasma.

An improved method for assaying paclitaxel in human plasma by high-performance liquid chromatography (HPLC) with UV detection at 227 nm has been developed by adapting previously reported sample preparation methods and chromatographic conditions to facilitate semi-automated sample cleanup using a column switching technique. Manual sample manipulations were limited to isolating the drug and internal standard from plasma (1.0 ml) by liquid-liquid extraction using tert-butyl methyl ether. The sample extract was initially loaded onto a short cartridge column containing a cyanopropyl stationary phase. During the predetermined time interval that the drug and internal standard eluted from the cartridge, 1.50-2.20 min, a fully automated 6-position switching valve was used to direct the effluent onto an octylsilica analytical column. The same mobile phase, composed of acetonitrile-methanol-ammonium acetate buffer (pH 5.0; 20 mM) (76:19:105, v/v/v) and delivered at flow rate of 1.0 ml/min, was used for both separations. The overall retention times of paclitaxel and the internal standard were 10.9 and 18.1 min, respectively. The analytical method was thoroughly validated for quantitating paclitaxel in plasma at concentrations ranging from 6 to 586 nM (5-500 ng/ml). The lowest concentration of paclitaxel measured with acceptable day-to-day accuracy (100.2%) and precision (RSD 11.7%, n = 21, 5 months) was 6 nM (5 ng/ml). The sensitivity and selectivity of the assay proved to be more than adequate for monitoring steady-state plasma concentrations of the drug when administered to cancer patients as a 96 h continuous intravenous infusion in combination with other anticancer agents, such as doxorubicin and topotecan. Moreover, the heart-cutting procedure prevented the problematic introduction of interfering nonpolar plasma components onto the analytical column, thereby enhancing sample throughput while decreasing the technical demands of the assay. The method was found to be extremely reproducible and robust during extended use for the routine analysis of plasma specimens acquired from several clinical trials.

Synergistic effect of heat-shock and UV-irradiation on mitosis and protein synthesis in G2-phase plasmodia of Physarum polycephalum Schw--reduction in UV-induced mitotic delay in a preheat-shocked system.

The synergistic effect of UV irradiation and heat-shock during the last 3 hr of G2 phase of the cell cycle in the plasmodia of P. polycephalum, in terms of mitotic delay and inhibition of protein synthesis, has been evaluated. The mitotic delay due to both perturbers coordinately increased closer to mitosis. Maximum mitotic delay was obtained in plasmodia heat-shocked after UV irradiation, indicating the presence in this system of either a heat-labile mitogenic substance which is comparatively less susceptible to UV or a substance which is made more susceptible to hyperthermia by UV. A protective role for heat-shock applied before irradiation has been observed in that, radiation-induced mitotic delay is significantly reduced in this combination. There was severe inhibition of translation in all the perturbed classes. Organelle level effects which are independent of major protein synthetic activities or different levels of heat-shock protein production could be the reason for the lack of correlation between percentage inhibition of general protein synthesis and the extent of mitotic delay with respect to the two double-perturbed systems.

Immunosuppression in cardiac transplantation: a new era in immunopharmacology.

During the past 5 years, there has been a concerted effort to identify new immunosuppressive agents for organ transplantation that have greater efficacy and fewer side effects than current therapies (corticosteroids, azathioprine, cyclosporine, anti-T cell antibodies). These new drugs can be generally classified according to their varying structures or mechanisms of action: xenobiotic molecules (FK506, rapamycin, cyclosporine analogues); antimetabolites (mycophenolate mofetil, mizoribine, brequinar sodium); and those agents with novel or incompletely defined mechanisms of action (leflunomide, 15-deoxyspergualin). Many of the newer agents offer better therapeutic indexes, and some even show promise in the treatment of two of the major obstacles currently facing cardiac allograft transplantation: antibody-mediated accelerated humoral rejection and chronic vascular rejection (also known as accelerated graft coronary artery disease). With the increasing shortage of donor organs in recent years, there has been a resurgence of interest in xenotransplantation; some of the new immunosuppressants demonstrate efficacy in prolonging xenograft survival. Most of these agents will probably find their niches as components of low-dose combination regimens designed to maximize effective immunosuppression and minimize drug toxicities and opportunistic infections.

Expression of plasmid-encoded aad in Clostridium acetobutylicum M5 restores vigorous butanol production.

Mutant M5 of Clostridium acetobutylicum ATCC 824, which produces neither butanol nor acetone and is deficient in butyraldehyde dehydrogenase (BYDH), acetoacetate decarboxylase, and acetoacetyl-coenzyme A:acetate/butyrate:coenzyme A-transferase activities, was transformed with plasmid pCAAD, which carries the gene aad (R. V. Nair, G. N. Bennett, and E. T. Papoutsakis, J. Bacteriol, 176:871-885, 1994). In batch fermentation studies, aad expression restored butanol formation (84 mM) in mutant M5 without any acetone formation or any significant increase in ethanol production. The corresponding protein (AAD) appeared as a ca. 96-kDa band in a denaturing protein gel. Expression of AAD in M5 resulted in restoration of BYDH activity and small increases in the activities of acetaldehyde dehydrogenase, butanol dehydrogenase, and ethanol dehydrogenase. These findings suggest that BYDH activity in C. acetobutylicum ATCC 824 resides largely in AAD, and that AAD's primary role is in the formation of butanol rather than of ethanol.

Novel route for the resolution of both enantiomers of dropropizine by using oxime esters and supported lipases of Pseudomonas cepacia.

Resolution of (R)- and (S)-dropropizine which is an antitussive and central sedative therapeutic agent in high optical and chemical yields was achieved by lipases of Pseudomonas cepacia supported on ceramic particles (lipase PS-C) and on diatomite (lipase PS-D) with oxime esters in organic solvents. The influence of several factors (lipase source, structural variations in oxime esters, the amount of lipase and its recyclability) on the enantioselectivity have been investigated. Different properties were used to describe the solvents, namely the hydrophobicity (quantified by log P) and the dielectic constant (epsilon). This enzymatic acylation using oxime esters was significant as only (S)-dropropizine and (R)-dropropizine monoacetate was obtained. (R)-Dropropizine monoacetate was chemically hydrolyzed to obtain (R)-dropropizine. The highest enantioselectivity was observed when O-acetyl benzophenone oxime was used. This enzymatic resolution provides a versatile method for getting the pure enantiomers of dropropizine by effectively optimizing the various reaction parameters.

Genetic analysis of resistance to whitebacked planthopper in twenty-one varieties of rice, Oryza sativa L.

The inheritance of resistance to whitebacked planthopper Sogatella furcifera (Horvath) was studied in 21 rice varieties. Reactions of F1; F2 and F3 progenies of the crosses of 21 resistant varieties with the susceptible variety 'TN 1' revealed that a single dominant gene governs resistance in 'Mushkan 41', 'Santhi', 'Siahnakidar 195', 'SM2-34', 'Tirisurkh 251', 'Zirijowaian 245', '18', '24A', '39', '76 S', '78', '180', '213 B', '267', '293', 'CI 6037-4', 'NP97', 'S39 JKW' and 'Bansphul'. In varieties '65' and '274 A', resistance is governed by one dominant and one recessive gene which segregate independently of each other. Tests for allelism with the Wbph 1 gene originally identified in 'N 22' revealed that the dominant gene present in all the test varieties is the same as Wbph 1. Further studies are required to determine the allelic relationships of the recessive gene found in varieties '65' and '274 A'.

The reaction of choline acetyltransferase with sulfhydryl reagents. Methoxycarbonyl-CoA disulfide as an active site-directed reagent.

The reaction of choline acetyltransferase with methoxycarbonyl alkyl disulfides leads to a progressive loss in enzyme activity as the size of the alkyl group increases from methyl to n-butyl. Reaction with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) or methoxycarbonyl coenzyme A (CoA) disulfide, leads to a total loss of enzyme activity. DTNB inactivation is biphasic (k1 = approximately 9 x 10(2) M-1 s-1, k2 = approximately 6 x 10(1) M-1 s-1) with the slow phase being diminished by acetyl-CoA. Methoxycarbonyl-CoA disulfide inactivation is also biphasic (k1 = approximately 2.1 x 10(3) M-1 s-1, k2 = approximately 6 x 10(1) M-1 s-1), with the rapid phase being diminished in the presence of acetyl-CoA. Inactivation by methoxycarbonyl methyl disulfide, ethyl disulfide, or hydroxyethyl disulfide, or by methyl methanethiosulfonate is not biphasic. Pretreatment of the enzyme with methyl methanethiosulfonate, which leads to a 25% loss in enzyme activity, abolishes the fast phase of DTNB inactivation, the slow phase of methoxycarbonyl-CoA disulfide inactivation, and any further inactivation by methoxycarbonyl ethyl disulfide. These results are interpreted to suggest that choline acetyltransferase contains two classes of reactive sulfhydryl groups, neither of which are required for enzyme activity.

Assay for the quinocarmycin analog DX-52-1 in human plasma using high-performance liquid chromatography with automated column switching and low wavelength ultraviolet detection.

The hydrocyanated derivative of the antitumor antibiotic quinocarmycin, DX-52-1 (I), exhibits impressive activity against human melanoma xenograft models in vivo. Phase I clinical trials to evaluate this compound as an antineoplastic agent have been initiated by the US National Cancer Institute. We have developed an HPLC assay for the determination of I in human plasma involving automated column switching and UV detection at 210 nm. The preparation of samples for chromatographic analysis entails the preliminary removal of plasma proteins by precipitation with acetonitrile, acidifying the clear supernatant to pH 4.5, then extracting twice with tert.-butyl methyl ether to recover the drug. A heart-cutting procedure employing two HPLC columns with contrasting retention characteristics under isocratic reversed-phase conditions was used to achieve the selectivity required for low wavelength UV detection of the analyte. The sample extract was initially loaded onto a column packed with a cyanopropyl stationary phase. During the predetermined time interval that I eluted from this column, a fully automated six-position switching valve was used to direct the effluent onto an octadecylsilane analytical column. The assay has been thoroughly validated with regard to linearity, inter- and intra-day accuracy and precision, recovery, selectivity and specificity. Using a sample volume of 1.0 ml, the lowest concentration of I quantified with acceptable day-to-day reproducibility was found to be 2.56 ng/ml (R.S.D. 18.9%, n=21, 4 months). This proved to be sufficiently sensitive for pharmacokinetic drug level monitoring in cancer patients treated with a 6-h continuous intravenous infusion of I, even at the starting dose of 3 mg/m2. The successful performance and reliability of the assay has been demonstrated through extensive application to the routine analysis of plasma specimens acquired during a phase I clinical trial of the drug.

Molecular characterization of an aldehyde/alcohol dehydrogenase gene from Clostridium acetobutylicum ATCC 824.

A gene (aad) coding for an aldehyde/alcohol dehydrogenase (AAD) was identified immediately upstream of the previously cloned ctfA (J. W. Cary, D. J. Petersen, E. T. Papoutsakis, and G. N. Bennett, Appl. Environ. Microbiol. 56:1576-1583, 1990) of Clostridium acetobutylicum ATCC 824 and sequenced. The 2,619-bp aad codes for a 96,517-Da protein. Primer extension analysis identified two transcriptional start sites 83 and 243 bp upstream of the aad start codon. The N-terminal section of AAD shows homology to aldehyde dehydrogenases of bacterial, fungal, mammalian, and plant origin, while the C-terminal section shows homology to alcohol dehydrogenases of bacterial (which includes three clostridial alcohol dehydrogenases) and yeast origin. AAD exhibits considerable amino acid homology (56% identity) over its entire sequence to the trifunctional protein encoded by adhE from Escherichia coli. Expression of aad from a plasmid in C. acetobutylicum showed that AAD, which appears as a approximately 96-kDa band in denaturing protein gels, provides elevated activities of NADH-dependent butanol dehydrogenase, NAD-dependent acetaldehyde dehydrogenase and butyraldehyde dehydrogenase, and a small increase in NADH-dependent ethanol dehydrogenase. A 957-bp open reading frame that could potentially encode a 36,704-Da protein was identified upstream of aad.

Covalent DNA adducts formed in mouse epidermis from dibenz[a,j]anthracene: evidence for the formation of polar adducts.

The formation of deoxyribonucleoside adducts in mouse epidermis has been examined following topical application of [3H]dibenz[a,j]anthracene (DB[a,j]A) or by 32P-postlabeling following topical application of unlabeled DB[a,j]A, DB[a,j]A trans-3,4-diol or the anti- or syn-3,4-diol 1,2-epoxides. A single topical application of [3H]DB[a,j]A at a dose of 400 nmol per mouse led to the formation of 11 detectable covalent DNA adducts. Seven of these DNA adducts were tentatively identified based on cochromatography with marker adducts using high-pressure liquid chromatography (HPLC). The presence of both deoxyguanosine (dGuo) as well as deoxyadenosine (dAdo) adducts formed from bay-region anti- and syn-3,4-diol 1,2-epoxides of DB[a,j]A was revealed. The major bay-region diol epoxide DNA adduct formed in mouse epidermis following topical application of [3H]DB[a,j]A was tentatively identified as the (4R,3S)-diol (2S,1R)-epoxide bound through trans addition of the exocyclic amino group of dGuo, although substantial amounts of the corresponding dAdo adduct were also detected. In addition, a K-region 5,6-oxide-dAdo adduct was tentatively identified in HPLC chromatograms based on cochromatography with an authentic marker adduct. 32P-Postlabeling analysis of DB[a,j]A-DNA adducts formed in mouse epidermis after topical application of unlabeled compound confirmed the presence of bay-region diol epoxide DNA adducts similar to those observed after application of [3H]DB[a,j]A. However, 32P-postlabeling analysis also revealed the presence of more polar covalent DNA adducts in epidermal DNA samples from DB[a,j]A-treated mice. These more polar DNA adducts represented a significant proportion of the 32P-labeled material recovered in HPLC chromatograms. While the exact nature of these adducts remains unknown at present, they had retention times identical to polar DNA adducts formed following topical application of DB[a,j]A trans-3,4-diol and may represent bis-dihydrodiol epoxide DNA adducts. The present results indicate that a rather broad spectrum of DNA adducts arises following topical application of DB[a,j]A to mouse epidermis.