Creation of a novel CRISPR-generated allele to express HA epitope-tagged C1QL1 and improved methods for its detection at synapses.

C1QL1 is expressed in a subset of cells in the brain and likely has pleiotropic functions, including the regulation of neuron-to-neuron synapses. Research progress on C1QL proteins has been slowed by a dearth of available antibodies. Therefore, we created a novel knock-in mouse line in which an HA-tag is inserted into the endogenous C1ql1 locus. We examined the entire brain, identifying previously unappreciated nuclei expressing C1QL1, presumably in neurons. By total numbers, however, the large majority of C1QL1-expressing cells are of the oligodendrocyte lineage. Subcellular immunolocalization of synaptic cleft proteins is challenging, so we developed a new protocol to improve signal at synapses. Lastly, we compared various anti-HA antibodies to assist future investigations using this and likely other HA epitope-tagged alleles.

Caldendrin Is a Repressor of PIEZO2 Channels and Touch Sensation in Mice.

The sense of touch is crucial for cognitive, emotional, and social development and relies on mechanically activated (MA) ion channels that transduce force into an electrical signal. Despite advances in the molecular characterization of these channels, the physiological factors that control their activity are poorly understood. Here, we used behavioral assays, electrophysiological recordings, and various mouse strains (males and females analyzed separately) to investigate the role of the calmodulin-like Ca2+ sensor, caldendrin, as a key regulator of MA channels and their roles in touch sensation. In mice lacking caldendrin (Cabp1 KO), heightened responses to tactile stimuli correlate with enlarged MA currents with lower mechanical thresholds in dorsal root ganglion neurons (DRGNs). The expression pattern of caldendrin in the DRG parallels that of the major MA channel required for touch sensation, PIEZO2. In transfected cells, caldendrin interacts with and inhibits the activity of PIEZO2 in a manner that requires an alternatively spliced sequence in the N-terminal domain of caldendrin. Moreover, targeted genetic deletion of caldendrin in Piezo2-expressing DRGNs phenocopies the tactile hypersensitivity of complete Cabp1 KO mice. We conclude that caldendrin is an endogenous repressor of PIEZO2 channels and their contributions to touch sensation in DRGNs.

A non-conducting role of the Cav1.4 Ca2+ channel drives homeostatic plasticity at the cone photoreceptor synapse.

In congenital stationary night blindness type 2 (CSNB2)-a disorder involving the Cav1.4 (L-type) Ca2+ channel-visual impairment is mild considering that Cav1.4 mediates synaptic release from rod and cone photoreceptors. Here, we addressed this conundrum using a Cav1.4 knockout (KO) mouse and a knock-in (G369i KI) mouse expressing a non-conducting Cav1.4. Surprisingly, Cav3 (T-type) Ca2+ currents were detected in cones of G369i KI mice and Cav1.4 KO mice but not in cones of wild-type mouse, ground squirrel, and macaque retina. Whereas Cav1.4 KO mice are blind, G369i KI mice exhibit normal photopic (i.e., cone-mediated) visual behavior. Cone synapses, which fail to form in Cav1.4 KO mice, are present, albeit enlarged, and with some errors in postsynaptic wiring in G369i KI mice. While Cav1.4 KO mice lack evidence of cone synaptic responses, electrophysiological recordings in G369i KI mice revealed nominal transmission from cones to horizontal cells and bipolar cells. In CSNB2, we propose that Cav3 channels maintain cone synaptic output provided that the nonconducting role of Cav1.4 in cone synaptogenesis remains intact. Our findings reveal an unexpected form of homeostatic plasticity that relies on a non-canonical role of an ion channel.

Development of genetic markers for reproductive management of toucans.

Retention of genetic diversity in successive generations is key to successful ex situ programs and will become increasingly important to restore wild populations of threatened animals. When animal genealogy is partly unknown or gaps exist in studbook records, the application of molecular resources facilitates informed breeding. Here, we apply molecular resources to an ex situ breeding population of toucans (Ramphastidae), a bird family zoos commonly maintain. Toucans face population declines from illegal poaching and habitat degradation. We developed novel microsatellite markers using blood samples from 15 Keel-billed Toucans (Ramphastos sulfuratus Lesson 1830). Parentage of two individuals was known a priori, but possible sibship among 13 putative founders-including the parents-was unknown. We compared available avian heterologous and novel microsatellite markers to recover known relationships and reconstruct sibship. Eight of 61 heterologous markers amplified consistently and were polymorphic, but less so than the 18 novel markers. Known sibship (and three sibling pairs whose relatedness was unknown a priori) and paternity-though not maternity except in one case-were well-recovered using both likelihood and pairwise relatedness methods, when incorporating novel but not heterologous markers. Zoo researchers seeking microsatellite primer sets for their breeding toucan populations will likely benefit from our heterologous markers, which can be leveraged both to assess relatedness and select breeding pairs. We recommend that zoo biologists rely on species-specific primers and not optimize heterologous primers for toucan species without molecular resources. We conclude with a brief discussion of modern genotyping methods of interest to zoo researchers.

Calcium Channels in Retinal Function and Disease.

Voltage-gated Ca2+ (Cav) channels play pivotal roles in regulating gene transcription, neuronal excitability, and neurotransmitter release. To meet the spatial and temporal demands of visual signaling, Cav channels exhibit unusual properties in the retina compared to their counterparts in other areas of the nervous system. In this article, we review current concepts regarding the specific subtypes of Cav channels expressed in the retina, their intrinsic properties and forms of modulation, and how their dysregulation could lead to retinal disease.

A systematic approach to transplanting non-resident, non-citizens in an established US pediatric lung transplant program.

Introduction: The Texas Children's Hospital Lung Transplant Program undertook consideration of its first non-resident, non-citizen for lung transplantation in 2011. Methods: Four referrals from the Royal Embassy of Saudi Arabia were received, and two patients were evaluated from 2011 to 2013. Results: After a suitable candidate and family was identified, the program adopted a systematic approach to ensure that all the necessary elements of pre-transplant care, informed consent, and post-transplant care could be effectively delivered. Conclusion: The use of hospital translation services and the development of a strong professional relationship with a well-trained pediatric respirologist in Saudi Arabia combined with an excellent early post-transplant clinical course provide lessons that may be of help to other transplant programs considering international patients as candidates for solid organ transplantation.

Epigenetic potential affects immune gene expression in house sparrows.

Epigenetic mechanisms may play a central role in mediating phenotypic plasticity, especially during range expansions, when populations face a suite of novel environmental conditions. Individuals may differ in their epigenetic potential (EP; their capacity for epigenetic modifications of gene expression), which may affect their ability to colonize new areas. One form of EP, the number of CpG sites, is higher in introduced house sparrows (Passer domesticus) than in native birds in the promoter region of a microbial surveillance gene, Toll-like Receptor 4 (TLR4), which may allow invading birds to fine-tune their immune responses to unfamiliar parasites. Here, we compared TLR4 gene expression from whole blood, liver and spleen in house sparrows with different EP, first challenging some birds with lipopolysaccharide (LPS), to increase gene expression by simulating a natural infection. We expected that high EP would predict high inducibility and reversibility of TLR4 expression in the blood of birds treated with LPS, but we did not make directional predictions regarding organs, as we could not repeatedly sample these tissues. We found that EP was predictive of TLR4 expression in all tissues. Birds with high EP expressed more TLR4 in the blood than individuals with low EP, regardless of treatment with LPS. Only females with high EP exhibited reversibility in gene expression. Further, the effect of EP varied between sexes and among tissues. Together, these data support EP as one regulator of TLR4 expression.

The complete mitochondrial genome of the oleaginous microalgae Ankistrodesmus falcatus strain UCP001 from the Peruvian Amazon.

Ankistrodesmus falcatus strain UCP001 is a native oleaginous microalgae isolated from the Peruvian Amazon basin. In this study we sequenced, de novo assembled, and functionally annotated the complete mitochondrial genome of the native oleaginous microalgae Ankistrodesmus falcatus strain UCP001 (Accesion number MT701044). This mitogenome is a typical circular double stranded DNA molecule of 41,048 bp in total length with G + C content of 37.4%. The mitogenome contains 49 genes, including 18 protein coding genes, 5 ribosomal (rRNA) genes and 26 transfer RNA (tRNA) genes. A phylogenetic analysis of 18 microalgae species indicated that Ankistrodesmus falcatus strain UCP001 was closely related to Ourococcus multisporus and Raphidocelis subcapitata. The complete mitochondrial genome sequence of Ankistrodesmus falcatus strain UCP001 enriches genomic resources of oleaginous native microalgae from the Peruvian Amazon for further basic and applied research.

Functional impact of a congenital stationary night blindness type 2 mutation depends on subunit composition of Cav1.4 Ca2+ channels.

Voltage-gated Cav1 and Cav2 Ca2+ channels are comprised of a pore-forming α1 subunit (Cav1.1-1.4, Cav2.1-2.3) and auxiliary ß (ß1-4) and α2δ (α2δ-1-4) subunits. The properties of these channels vary with distinct combinations of Cav subunits and alternative splicing of the encoding transcripts. Therefore, the impact of disease-causing mutations affecting these channels may depend on the identities of Cav subunits and splice variants. Here, we analyzed the effects of a congenital stationary night blindness type 2 (CSNB2)-causing mutation, I745T (IT), in Cav1.4 channels typical of those in human retina: Cav1.4 splice variants with or without exon 47 (Cav1.4+ex47 and Cav1.4Δex47, respectively), and the auxiliary subunits, ß2X13 and α2δ-4. We find that IT caused both Cav1.4 splice variants to activate at significantly more negative voltages and with slower deactivation kinetics than the corresponding WT channels. These effects of the IT mutation, along with unexpected alterations in ion selectivity, were generally larger in channels lacking exon 47. The weaker ion selectivity caused by IT led to hyperpolarizing shifts in the reversal potential and large outward currents that were evident in channels containing the auxiliary subunits ß2X13 and α2δ-4 but not in those with ß2A and α2δ-1. We conclude that the IT mutation stabilizes channel opening and alters ion selectivity of Cav1.4 in a manner that is strengthened by exclusion of exon 47 and inclusion of ß2X13 and α2δ-4. Our results reveal complex actions of IT in modifying the properties of Cav1.4 channels, which may influence the pathological consequences of this mutation in retinal photoreceptors.

A dual role for Cav1.4 Ca2+ channels in the molecular and structural organization of the rod photoreceptor synapse.

Synapses are fundamental information processing units that rely on voltage-gated Ca2+ (Cav) channels to trigger Ca2+-dependent neurotransmitter release. Cav channels also play Ca2+-independent roles in other biological contexts, but whether they do so in axon terminals is unknown. Here, we addressed this unknown with respect to the requirement for Cav1.4 L-type channels for the formation of rod photoreceptor synapses in the retina. Using a mouse strain expressing a non-conducting mutant form of Cav1.4, we report that the Cav1.4 protein, but not its Ca2+ conductance, is required for the molecular assembly of rod synapses; however, Cav1.4 Ca2+ signals are needed for the appropriate recruitment of postsynaptic partners. Our results support a model in which presynaptic Cav channels serve both as organizers of synaptic building blocks and as sources of Ca2+ ions in building the first synapse of the visual pathway and perhaps more broadly in the nervous system.

Nutritional evaluation and human health-promoting potential of compounds biosynthesized by native microalgae from the Peruvian Amazon.

A plausible strategy to mitigate socioeconomic problems in the Peruvian Amazon is through the sustainable exploitation of biodiversity resources, such as native microalgae. Several studies worldwide affirm that these microorganisms are excellent sources of higher value products for human nutrition and possess health-promoting biochemicals, but these attributes are unknown for the native microalgae of Peru. Therefore, the aim of this investigation was to evaluate the nutritional and human health-promoting potential of compounds biosynthesized by native microalgae from the Peruvian Amazon. Ten native microalgae strains of the groups cyanobacteria and chlorophyta were cultured in BG-11 medium and their biomass harvested and dried. Standardized methods were then used to determine proximate composition, fatty acids and amino acids composition, antioxidant activity, and total phenolic content. All ten microalgae strains produce primary nutrients, the entire spectrum of essential amino acids, essential fatty acids, and 3 of the 10 microalgae strains produced eisosapentaenoic acid. Additionally, all microalgae strains exhibited antioxidant activities and contained phenolic compounds. In conclusion, native microalgae strains from the Peruvian Amazon analyzed in this study possess the ability to biosynthesize and accumulate several nutrients and compounds with human health-promoting potential.

A single dose of zoledronate preserves bone mineral density for up to 2 years after a second course of romosozumab.

This phase 2 study evaluated the efficacy and safety of transitioning to zoledronate following romosozumab treatment in postmenopausal women with low bone mass. A single dose of 5 mg zoledronate generally maintained the robust BMD gains accrued with romosozumab treatment and was well tolerated. INTRODUCTION: Follow-on therapy with an antiresorptive agent is necessary to maintain the skeletal benefits of romosozumab therapy. We evaluated the use of zoledronate following romosozumab treatment. METHODS: This phase 2, dose-finding study enrolled postmenopausal women with low bone mineral density (BMD). Subjects who received various romosozumab doses or placebo from months 0-24 were rerandomized to denosumab (60 mg SC Q6M) or placebo for 12 months, followed by open-label romosozumab (210 mg QM) for 12 months. At month 48, subjects who had received active treatment for 48 months were assigned to no further active treatment and all other subjects were assigned to zoledronate 5 mg IV. Efficacy (BMD, P1NP, and ß-CTX) and safety were evaluated for 24 months, up to month 72. RESULTS: A total of 141 subjects entered the month 48-72 period, with 51 in the no further active treatment group and 90 in the zoledronate group. In subjects receiving no further active treatment, lumbar spine (LS) BMD decreased by 10.8% from months 48-72 but remained 4.2% above the original baseline. In subjects receiving zoledronate, LS BMD was maintained (percentage changes: - 0.8% from months 48-72; 12.8% from months 0-72). Similar patterns were observed for proximal femur BMD in both groups. With no further active treatment, P1NP and ß-CTX decreased but remained above baseline at month 72. Following zoledronate, P1NP and ß-CTX levels initially decreased but approached baseline by month 72. No new safety signals were observed. CONCLUSION: A zoledronate follow-on regimen can maintain robust BMD gains achieved with romosozumab treatment.

Dataset of de novo assembly and functional annotation of the transcriptomes of three native oleaginous microalgae from the Peruvian Amazon.

Microalgae are photosynthetic organisms with cosmopolitan distribution (i.e., marine, freshwater and terrestrial habitats) and possess a great diversity of species [1] and consequently an immense variation in biochemical compositions [2]. To date genomic information is available mainly from the model green microalga Chlamydomonas reinhardtii[3]. Here we provide the dataset of a de novo assembly and functional annotation of the transcriptomes of three native oleaginous microalgae from the Peruvian Amazon. Native oleaginous microalgae species Ankistrodesmus sp., Chlorella sp., and Scenedesmus sp. were cultured in triplicate using Chu-10 medium with or without a source of nitrate (NaNO3). Total RNA was purified, the cDNA libraries were constructed and sequenced as paired-end reads on an Illumina HiSeq™2500 platform. Transcriptomes were de novo assembled using Trinity v2.9.1. A total of 48,554 transcripts (range from 250 to 7966 bp; N50 = 1047) for Ankistrodesmus sp., 108,126 transcripts (range from 250 to 8160 bp; N50 = 1090) for Chlorella sp., and 77,689 transcripts (range from 250 to 8481 bp; N50 = 1281) for Scenedesmus sp. were de novo assembled. Completeness of the assembled transcriptomes were evaluated with the Benchmarking Universal Single-Copy Orthologs (BUSCO) software v2/v3. Functional annotation of the assembled transcriptomes was conducted with TransDecoder v3.0.1 and the web-based platforms Kyoto Encyclopedia of Genes and Genomes (KEGG) Automatic Annotation Server (KAAS) and FunctionAnnotator. The raw reads were deposited into NCBI and are accessible via BioProject accession number PRJNA628966 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA628966) and Sequence Read Archive (SRA) with accession numbers: SRX8295665 (https://www.ncbi.nlm.nih.gov/sra/SRX8295665), SRX8295666 (https://www.ncbi.nlm.nih.gov/sra/SRX8295666), SRX8295667 (https://www.ncbi.nlm.nih.gov/sra/SRX8295667), SRX8295668 (https://www.ncbi.nlm.nih.gov/sra/SRX8295668), SRX8295669 (https://www.ncbi.nlm.nih.gov/sra/SRX8295669), and SRX8295670 (https://www.ncbi.nlm.nih.gov/sra/SRX8295670). Additionally, transcriptome shotgun assembly sequences and functional annotations are available via Discover Mendeley Data (https://data.mendeley.com/datasets/47wdjmw9xr/1).

Dataset of de novo assembly and functional annotation of the transcriptome during germination and initial growth of seedlings of Myrciaria Dubia "camu-camu".

Myrciaria dubia "camu-camu" is a native shrub of the Amazon that is commonly found in areas that are flooded for three to four months during the annual hydrological cycle. This plant species is exceptional for its capacity to biosynthesize and accumulate important quantities of a variety of health-promoting phytochemicals, especially vitamin C [1], yet few genomic resources are available [2]. Here we provide the dataset of a de novo assembly and functional annotation of the transcriptome from a pool of samples obtained from seeds during the germination process and seedlings during the initial growth (until one month after germination). Total RNA/mRNA was purified from different types of plant materials (i.e., imbibited seeds, germinated seeds, and seedlings of one, two, three, and four weeks old), pooled in equimolar ratio to generate the cDNA library and RNA paired-end sequencing was conducted on an Illumina HiSeq™2500 platform. The transcriptome was de novo assembled using Trinity v2.9.1 and SuperTranscripts v2.9.1. A total of 21,161 transcripts were assembled ranging in size from 500 to 10,001 bp with a N50 value of 1,485 bp. Completeness of the assembly dataset was assessed using the Benchmarking Universal Single-Copy Orthologs (BUSCO) software v2/v3. Finally, the assembled transcripts were functionally annotated using TransDecoder v3.0.1 and the web-based platforms Kyoto Encyclopedia of Genes and Genomes (KEGG) Automatic Annotation Server (KAAS), and FunctionAnnotator. The raw reads were deposited into NCBI and are accessible via BioProject accession number PRJNA615000 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA615000) and Sequence Read Archive (SRA) with accession number SRX7990430 (https://www.ncbi.nlm.nih.gov/sra/SRX7990430). Additionally, transcriptome shotgun assembly sequences and functional annotations are available via Discover Mendeley Data (https://data.mendeley.com/datasets/2csj3h29fr/1).

Epigenetic Potential in Native and Introduced Populations of House Sparrows (Passer domesticus).

Epigenetic potential, defined as the capacity for epigenetically-mediated phenotypic plasticity, may play an important role during range expansions. During range expansions, populations may encounter relatively novel challenges while experiencing lower genetic diversity. Phenotypic plasticity via epigenetic potential might be selectively advantageous at the time of initial introduction or during spread into new areas, enabling introduced organisms to cope rapidly with novel challenges. Here, we asked whether one form of epigenetic potential (i.e., the abundance of CpG sites) in three microbial surveillance genes: Toll-like receptors (TLRs) 1B (TLR1B), 2A (TLR2A), and 4 (TLR4) varied between native and introduced house sparrows (Passer domesticus). Using an opportunistic approach based on samples collected from sparrow populations around the world, we found that introduced birds had more CpG sites in TLR2A and TLR4, but not TLR1B, than native ones. Introduced birds also lost more CpG sites in TLR1B, gained more CpG sites in TLR2A, and lost fewer CpG sites in TLR4 compared to native birds. These results were not driven by differences in genetic diversity or population genetic structure, and many CpG sites fell within predicted transcription factor binding sites (TFBS), with losses and gains of CpG sites altering predicted TFBS. Although we lacked statistical power to conduct the most rigorous possible analyses, these results suggest that epigenetic potential may play a role in house sparrow range expansions, but additional work will be critical to elucidating how epigenetic potential affects gene expression and hence phenotypic plasticity at the individual, population, and species levels.

Metagenomic 16S rDNA amplicon data on bacterial diversity profiling and its predicted metabolic functions of varillales in Allpahuayo-Mishana National Reserve.

The white-sands forests or varillales of the Peruvian Amazon are characterized by their distinct physical characteristics, patchy distribution, and endemism [1, 2]. Much research has been conducted on the specialized plant and animal communities that inhabit these ecosystems, yet their soil microbiomes have yet to be studied. Here we provide metagenomic 16S rDNA amplicon data of soil microbiomes from three types of varillales in Allpahuayo-Mishana National Reserve near Iquitos, Peru. Composite soil samples were collected from very low varillal, high-dry varillal, and high-wet varillal. Purified metagenomic DNA was used to prepare and sequence 16S rDNA metagenomic libraries on the Illumina MiqSeq platform. Raw paired-endsequences were analyzed using the Metagenomics RAST server (MG-RAST) and Parallel-Meta3 software and revealed the existence of a high percentage of undiscovered sequences, potentially indicating specialized bacterial communities in these forests. Also, were predicted several metabolic functions in this dataset. The raw sequence data in fastq format is available in the public repository Discover Mendeley Data (https://data.mendeley.com/datasets/syktzxcnp6/2). Also, is available at NCBI's Sequence Read Archive (SRA) with accession numbers SRX7891206 (very low varillal), SRX7891207 (high-dry varillal), and SRX7891208 (high-wet varillal).

Microsatellite primer development in elasmobranchs using next generation sequencing of enriched libraries.

Microsatellites are useful in studies of population genetics, sibship, and parentage. Here, we screened for microsatellites from multiple elasmobranch genomic libraries using an enrichment protocol followed by sequencing on an Illumina platform. We concurrently screened five and then nine genomes and describe the number of potential loci from each respective round of sequencing. To validate the efficacy of the protocol, we developed and tested primers for the pelagic thresher shark, Alopias pelagicus. The method described here is a cost-effective protocol to increase the pool of potential useful loci and allows the concurrent screening of multiple libraries.

Romosozumab or alendronate for fracture prevention in East Asian patients: a subanalysis of the phase III, randomized ARCH study.

Romosozumab, a sclerostin antibody, exerts dual effect to increase bone formation and decrease bone resorption. Among high-risk postmenopausal East Asian women, romosozumab followed by alendronate was associated with lower incidences of fractures vs alendronate alone. Romosozumab demonstrates potential to address an unmet need in osteoporosis management in Asia. INTRODUCTION: Romosozumab, a sclerostin antibody, exerts dual effect to increase bone formation and decrease bone resorption. The global ARCH study demonstrated superiority of romosozumab followed by alendronate in reducing fracture risk in high-risk postmenopausal osteoporotic women vs alendronate alone. We report outcomes among ARCH East Asian patients. METHODS: In ARCH, 4093 postmenopausal osteoporotic women with fragility fracture were randomized 1:1 to monthly romosozumab 210 mg or weekly alendronate 70 mg for 12 months, both followed by open-label alendronate. Primary endpoints were incidence of new vertebral fracture (VF) at 24 months and clinical fracture at primary analysis (confirmed fractures in ≥ 330 patients and all patients had opportunity to attend month 24 visit). This post hoc analysis was not powered to detect fracture-rate differences. RESULTS: This analysis included 275 patients from Hong Kong, Korea, and Taiwan. Romosozumab followed by alendronate reduced risk of new VFs at 24 months by 60% (P = 0.11) and clinical fractures at primary analysis by 44% (P = 0.15) vs alendronate alone. Romosozumab followed by alendronate significantly increased mean bone mineral density at 24 months from baseline by a further 9.0%, 3.3%, and 3.0% at the lumbar spine, total hip, and femoral neck vs alendronate alone. Adverse event (AE) rates, including positively adjudicated serious cardiovascular AEs (1.6% vs 1.4% at 12 months for romosozumab vs alendronate), were similar across treatment groups. CONCLUSIONS: Consistent with the global analysis, romosozumab followed by alendronate was associated with lower incidences of new vertebral, clinical, non-vertebral, and hip fractures vs alendronate alone among East Asian patients.

Bone mineral density gains with a second 12-month course of romosozumab therapy following placebo or denosumab.

Romosozumab is a therapy that stimulates bone formation and reduces bone resorption. In this study of postmenopausal women with low BMD, a second course of romosozumab following a period off treatment or on denosumab increased or maintained BMD, respectively, and was well tolerated, providing insight into treatment sequence options. INTRODUCTION: In patients with high fracture risk, therapies that stimulate bone formation provide rapid BMD gains; currently available agents, parathyroid hormone receptor agonists, are limited to a 2-year lifetime exposure and generally used for a single treatment course. However, for long-term osteoporosis management, a second treatment course may be appropriate. Romosozumab, a therapy with the dual effect of increasing bone formation and decreasing bone resorption, reduces fracture risk within 12 months. Here, we report efficacy and safety of a second romosozumab course. METHODS: In this phase 2, dose-finding study, postmenopausal women with low bone mass (T-score ≤ - 2.0 and ≥ - 3.5) received romosozumab or placebo (month 0-24) followed by placebo or denosumab (month 24-36); participants then received a year of romosozumab (month 36-48). RESULTS: Of 167 participants who entered the month 36-48 period, 35 had been initially randomized to romosozumab 210 mg monthly. In participants who received romosozumab 210 mg monthly followed by placebo, a second romosozumab course (n = 19) increased BMD by amounts similar to their initial treatment (month 0-12) at the lumbar spine (12.4%; 12.0%, respectively) and total hip (6.0%; 5.5%, respectively). Following denosumab, a second romosozumab course (n = 16) increased BMD at the lumbar spine (2.3%) and maintained BMD at the total hip. Safety profiles were similar between first and second romosozumab courses. CONCLUSIONS: After 12 months off-treatment, a second romosozumab course again led to rapid and large BMD gains. Following denosumab, BMD gains with romosozumab were smaller than with initial treatment. No new safety findings were observed during the second course.

In Silico Characterization and Expression Analysis Of The Alpha Subunit Of The Heteromeric Acetyl-Coenzyme A Carboxylase From Two Microalgae / Caracterización in silico y análisis de la expresión de la subunidad alfa de la acetil-coenzima a carboxilasa heteromérica de dos microalgas

RESUMEN Las microalgas son microorganismos fotosintéticos con gran potencial para abastecer las demandas energéticas mundiales. Sin embargo, los limitados conocimientos que se tienen de estos organismos, en particular a nivel molecular de los procesos metabólicos, han limitado su uso con estos propósitos. En esta investigación se ha realizado el análisis in silico de la subunidad alfa de la acetil-Coenzima A carboxilasa heteromérica (αACCasa), una enzima clave en la biosíntesis de lípidos de las microalgas Chlorella sp. y Scenedesmus sp. Asimismo, se ha medido la expresión de este gen en ambas especies cultivadas en medios deficientes de nitrógeno. Los resultados indican que la αACCasa muestra conservación estructural y funcional en ambas especies de microalgas y su mayor similitud genética con otras especies de microalgas. Asimismo, se ha mostrado que el nivel de expresión del gen se incrementa significativamente cuando las microalgas son cultivadas en ausencia de nitrógeno, lo cual se relaciona a su vez con una mayor acumulación de lípidos microalgales. En conclusión, el análisis in silico de la αACCasa de Chlorella sp. y Scenedesmus sp. presentan características estructurales, funcionales y evolutivas muy similares con otras especies de microalgas y plantas. Asimismo, el estudio revela que en ambas especies el gen se sobreexpresa cuando las microalgas son sometidas a estrés por deficiencia de nitrógeno, el cual se relaciona significativamente con la acumulación de lípidos totales en estas células.

ABSTRACT Microalgae are photosynthetic microorganisms with great potential to supply the world's energy demands. However, the limited knowledge of these organisms, particularly at the molecular level of metabolic processes, has limited their use to these purposes. In this investigation, the in silico analysis of the alpha subunit of the heteromeric acetyl-coenzyme A carboxylase (αACCase), a key enzyme in lipid biosynthesis of microalgae Chlorella sp. and Scenedesmus sp. was carried out. Also, the expression of this gene has been measured in both species cultivated in nitrogen-depleted media. Results indicate that αACCase shows structural and functional conservation in both species of microalgae and their greater genetic similarity with other species of microalgae. Also, it has been shown that the expression levels of this gene are significantly increased when the microalgae are cultured in the absence of nitrogen, which in turn is related to a greater accumulation of microalgal lipids. In conclusion, the in silico analysis of the Chlorella sp. and Scenedesmus sp. αACCase reveals structural, functional and evolutionary characteristics very similar to other microalgae and plant species. Also, the study reveals that in both species the gene is overexpressed when microalgae are subjected to nitrogen deficiency stress, which is significantly related to total lipids accumulation in these cells.