Multiple deficiencies of mitochondrial DNA- and nuclear-encoded subunits of respiratory NADH dehydrogenase detected with peptide- and subunit-specific antibodies in mitochondrial myopathies

AbstractAntibodies have been raised against synthetic peptides corresponding to several computer-predicted epitopes of three mtDNA-encoded subunits, ND4, ND5 and ND6, of the human respiratory chain NADH dehydrogenase (Complex I). Antibodies were characterized by a sensitive immunoblotting assay using proteins from human skeletal muscle mitochondria and by immunoprecipitation of radio-labeled HeLa cell mitochondrial translation products. Only antibodies against two of six selected peptides of the ND4 subunit, i.e., the C-terminal peptide and an internal peptide close to the C-terminus, reacted in both assays with the subunit. Antibodies raised against an internal peptide close to the N-terminus of the ND5 subunit and antibodies raised against an internal epitope of the ND6 subunit also reacted in both the immunoblotting and immunoprecipitation assays. The antibodies described above and other Complex I subunit- or holoenzyme-specific antibodies were used to investigate the subunit deficiencies of the respiratory NADH dehydrogenase in the skeletal muscle of patients affected by mitochondrial myopathies associated with Complex I defects. The reduction in enzyme activity correlated in an immunoblot assay with a decrease of four mtDNA-encoded subunits of the enzyme, as well as with a decrease of other subunits of Complex I encoded in the nDNA. The present work provides the first evidence of a decrease in NADH dehydrogenase subunits encoded in the mitochondrial genome in myopathy patients

In situ monitoring of GaSb, GaInAsSb, and AlGaAsSb

Suitability of silicon photodiode detector arrays for monitoring the spectral reflectance during epitaxial growths of GaSb, AlGaAsSb, and GaInAsSb, which have cutoff wavelengths of 1.7, 1.2, and 2.3 {micro}m, respectively, is demonstrated. These alloys were grown lattice matched to GaSb in a vertical rotating-disk reactor, which was modified to accommodate near normal reflectance without affecting epilayer uniformity. By using a virtual interface model, the growth rate and complex refractive index at the growth temperature are extracted for these alloys over the 600 to 950 nm spectral range. Excellent agreement is obtained between the extracted growth rate and that determined by ex-situ measurement. Optical constants are compared to theoretical predictions based on an existing dielectric function model for these materials. Furthermore, quantitative analysis of the entire reflectance spectrum yields valuable information on the approximate thickness of overlayers on the pregrowth substrate

Optimal low-thrust trajectories to asteroids through an algorithm based on differential dynamic programming

In this paper an optimisation algorithm based on Differential Dynamic Programming is applied to the design of rendezvous and fly-by trajectories to near Earth objects. Differential dynamic programming is a successive approximation technique that computes a feedback control law in correspondence of a fixed number of decision times. In this way the high dimensional problem characteristic of low-thrust optimisation is reduced into a series of small dimensional problems. The proposed method exploits the stage-wise approach to incorporate an adaptive refinement of the discretisation mesh within the optimisation process. A particular interpolation technique was used to preserve the feedback nature of the control law, thus improving robustness against some approximation errors introduced during the adaptation process. The algorithm implements global variations of the control law, which ensure a further increase in robustness. The results presented show how the proposed approach is capable of fully exploiting the multi-body dynamics of the problem; in fact, in one of the study cases, a fly-by of the Earth is scheduled, which was not included in the first guess solution

The CXCL12/CXCR4 signaling axis retains neutrophils at inflammatory sites in zebrafish

The inappropriate retention of neutrophils at inflammatory sites is a major driver of the excessive tissue damage characteristic of respiratory inflammatory diseases including COPD, ARDS, and cystic fibrosis. The molecular programmes which orchestrate neutrophil recruitment to inflammatory sites through chemotactic guidance have been well-studied. However, how neutrophil sensitivity to these cues is modulated during inflammation resolution is not understood. The identification of neutrophil reverse migration as a mechanism of inflammation resolution and the ability to modulate this therapeutically has identified a new target to treat inflammatory disease. Here we investigate the role of the CXCL12/CXCR4 signaling axis in modulating neutrophil retention at inflammatory sites. We used an in vivo tissue injury model to study neutrophilic inflammation using transgenic zebrafish larvae. Expression of cxcl12a and cxcr4b during the tissue damage response was assessed using in situ hybridization and analysis of RNA sequencing data. CRISPR/Cas9 was used to knockdown cxcl12a and cxcr4b in zebrafish larvae. The CXCR4 antagonist AMD3100 was used to block the Cxcl12/Cxcr4 signaling axis pharmacologically. We identified that cxcr4b and cxcl12a are expressed at the wound site in zebrafish larvae during the inflammatory response. Following tail-fin transection, removal of neutrophils from inflammatory sites is significantly increased in cxcr4b and cxcl12a CRISPR knockdown larvae. Pharmacological inhibition of the Cxcl12/Cxcr4 signaling axis accelerated resolution of the neutrophil component of inflammation, an effect caused by an increase in neutrophil reverse migration. The findings of this study suggest that CXCR4/CXCL12 signaling may play an important role in neutrophil retention at inflammatory sites, identifying a potential new target for the therapeutic removal of neutrophils from the lung in chronic inflammatory disease

Evolution of HCl Concentrations in the Lower Stratosphere from 1991 to 1996 Following the Eruption of Mt. Pinatubo

Geophysical Research Letters, Vol. 25, No. 7, pp. 995-998, April 1, 1998.In situ measurements of hydrochloric acid in the lower stratosphere reveal that its mean abundance relative to that of total inorganic chlorine..

POU5F1 (OCT3/4) identifies cells with pluripotent potential in human germ cell tumors

Human germ cell tumors (GCTs) may have variable histology and clinical behavior, depending on factors such as sex of the patient, age at clinical diagnosis, and anatomical site of the tumor. Some types of GCT, i.e., the seminomas/germinomas/dysgerminomas and embryonal carcinomas (the stem cell component of nonseminomas), have pluripotent potential, which is demonstrated by their capacity to differentiate into somatic and/or extraembryonic elements. Although embryonal carcinoma cells are intrinsically pluripotent, seminoma/germinoma/dysgerminoma cells, as well as their precursor carcinoma in situ/gonadoblastoma cells, have the phenotype of early germ cells that can be activated to pluripotency. The other types of GCT (teratomas and yolk sac tumors of infants and newborn, dermoid cyst of the ovary, and spermatocytic seminoma of elderly) are composed of (fully) differentiated tissues and lack the appearance of undifferentiated and pluripotent stem cells. OCT3/4, a transcription factor also known as OTF3 and POU5F1, is involved in regulation of pluripotency during normal development and is detectable in embryonic stem and germ cells. We analyzed the presence of POU5F1 in GCT and other tumor types using immunohistochemistry. The protein was consistently detected in carcinoma in situ/gonadoblasto

Lipophilic nalmefene prodrugs to achieve a one-month sustained release

Nalmefene is an opioid antagonist which as a once-a-day tablet formulation has recently been approved for reducing ethanol intake in alcoholic subjects. In order to address the compliance issue in this patient population, a number of potential nalmefene prodrugs were synthesized with the aim of providing a formulation that could provide plasma drug concentrations in the region of 0.5–1.0 ng/mL for a one-month period when dosed intramuscular to dogs or minipigs. In an initial series of studies, three different lipophilic nalmefene derivatives were evaluated: the palmitate (C16), the octadecyl glutarate diester (C18-C5) and the decyl carbamate (CB10). They were administered intramuscularly to dogs in a sesame oil solution at a dose of 1 mg-eq. nalmefene/kg. The decyl carbamate was released relatively quickly from the oil depot and its carbamate bond was too stable to be used as a prodrug. The other two derivatives delivered a fairly constant level of 0.2–0.3 ng nalmefene/mL plasma for one month and since there was no significant difference between these two, the less complex palmitate monoester was chosen to demonstrate that dog plasma nalmefene concentrations were dose-dependent at 1, 5 and 20 mg-eq. nalmefene/kg. In a second set of experiments, the effect of the chain length of the fatty acid monoester promoieties was examined. The increasingly lipophilic octanoate (C8), decanoate (C10) and dodecanoate (C12) derivatives were evaluated in dogs and in minipigs, at a dose of 5 mg-eq. nalmefene/kg and plasma nalmefene concentrations were measured over a four-week period. The pharmacokinetic profiles were very similar in both species with Cmax decreasing and Tmax increasing with increasing fatty acid chain length and the target plasma concentrations (0.5–1.0 ng/mL over a month-long period) were achieved with the dodecanoate (C12) prodrug. These data therefore demonstrate that sustained plasma nalmefene concentrations can be achieved in both dog and minipig using nalmefene prodrugs and that the pharmacokinetic profile of nalmefene can be tuned by varying the length of the alkyl group