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Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a protease that regulates low density lipoprotein receptor (LDLR) protein levels. The mechanisms of this action, however, remain to be defined. We show here that recombinant human PCSK9 expressed in HEK293 cells was readily secreted into the medium, with the prosegment associated with the C-terminal domain. Secreted PCSK9 mediated cell surface LDLR degradation in a concentration- and time-dependent manner when added to HEK293 cells. Accordingly, cellular LDL uptake was significantly reduced as well. When infused directly into C57B6 mice, purified human PCSK9 substantially reduced hepatic LDLR protein levels and resulted in increased plasma LDL cholesterol. When added to culture medium, fluorescently labeled PCSK9 was endocytosed and displayed endosomal-lysosomal intracellular localization in HepG2 cells, as was demonstrated by colocalization with DiI-LDL. PCSK9 endocytosis was mediated by LDLR as LDLR deficiency (hepatocytes from LDLR null mice), or RNA interference-mediated knockdown of LDLR markedly reduced PCSK9 endocytosis. In addition, RNA interference knockdown of the autosomal recessive hypercholesterolemia (ARH) gene product also significantly reduced PCSK9 endocytosis. Biochemical analysis revealed that the LDLR extracellular domain interacted directly with secreted PCSK9; thus, overexpression of the LDLR extracellular domain was able to attenuate the reduction of cell surface LDLR levels by secreted PCSK9. Together, these results reveal that secreted PCSK9 retains biological activity, is able to bind directly to the LDLR extracellular domain, and undergoes LDLR-ARH-mediated endocytosis, leading to accelerated intracellular degradation of the LDLR.

Hypercholesterolemia is a major risk factor for coronary artery disease. Oxysterols are known to inhibit cholesterol biosynthesis and have been explored as potential antihypercholesterolemic agents. The ability of 3β-hydroxy-5α-cholest-8(14)-en-15-one (15-ketosterol) to lower non-HDL cholesterol has been demonstrated in rodent and primate models, but the mechanisms of action remain poorly understood. Here we show in a coactivator recruitment assay and cotransfection assays that the 15-ketosterol is a partial agonist for liver X receptor-α and -β (LXRα and LXRβ). The binding affinity for the LXRs was comparable to those of native oxysterols. In a macrophage cell line of human origin, the 15-ketosterol elevated ATP binding cassette transporter ABCA1 mRNA in a concentration-dependent fashion with a potency similar to those of other oxysterols. We further found that in human embryonic kidney HEK 293 cells, the 15-ketosterol suppressed sterol-responsive element binding protein processing activity and thus inhibited mRNA expression of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, LDL receptor, and PCSK9. Our data thus provide a molecular basis for the hypocholesterolemic activity of the 15-ketosterol and further suggest its potential antiatherosclerotic benefit as an LXR agonist.

Estrogen replacement therapy in women decreases hepatic lipase (HL) activity, which may account for the associated increase in HDL cholesterol. To investigate whether estrogen decreases HL transcription, transient cotransfection assays with HL promoter and estrogen receptor-α (ERα) expression constructs were performed in HepG2 cells. 17β-estradiol (E2) decreased transcription driven by the −1557/+41 human HL promoter by up to 50% at 10−7 M. Mutation of ERα by deletion of its transactivation domains or ligand-binding domain eliminated E2-induced repression of the promoter, whereas deletion of the DNA-binding domain of ERα resulted in a 7-fold activation by E2. The E2-induced repression was maintained after mutation of a potential estrogen-response element in the promoter. The region of estrogen responsiveness was localized to −1557/ −1175 of the HL promoter by deletion analysis. Mutation of an AP-1 site at −1493 resulted in a partial loss of E2-induced repression, similar to that caused by deletion of nucleotides −1557 to −1366. Gel shift assays with nuclear extracts from E2-treated HepG2 cells stably expressing ERα demonstrated an increase in binding to an AP-1 consensus oligonucleotide. The AP-1 activator, phorbol 12-myristate 13-acetate, inhibited the HL promoter by greater than 50%. Collectively, the data suggest that estrogen represses the transcription of the HL gene, possibly through an AP-1 pathway. —Jones, D. R., R. J. Schmidt, R. T. Pickard, P. S. Foxworthy, and P. I. Eacho. Estrogen receptor-mediated repression of human hepatic lipase gene transcription.

Social structure is proposed to influence the transmission of both directly and environmentally transmitted infectious agents. However in natural populations, many other factors also influence transmission, including variation in individual susceptibility and aspects of the environment that promote or inhibit exposure to infection. We used a population genetic approach to investigate the effects of social structure, environment, and host traits on the transmission of Escherichia coli infecting two populations of wild elephants: one in Amboseli National Park and another in Samburu National Reserve, Kenya. If E. coli transmission is strongly influenced by elephant social structure, E. coli infecting elephants from the same social group should be genetically more similar than E. coli sampled from members of different social groups. However, we found no support for this prediction. Instead, E. coli was panmictic across social groups, and transmission patterns were largely dominated by habitat and host traits. For instance, habitat overlap between elephant social groups predicted E. coli genetic similarity, but only in the relatively drier habitat of Samburu, and not in Amboseli, where the habitat contains large, permanent swamps. In terms of host traits, adult males were infected with more diverse haplotypes, and males were slightly more likely to harbor strains with higher pathogenic potential, as compared to adult females. In addition, elephants from similar birth cohorts were infected with genetically more similar E. coli than elephants more disparate in age. This age-structured transmission may be driven by temporal shifts in genetic structure of E. coli in the environment and the effects of age on bacterial colonization. Together, our results support the idea that, in elephants, social structure often will not exhibit strong effects on the transmission of generalist, fecal-oral transmitted bacteria. We discuss our results in the context of social, environmental, and host-related factors that influence transmission patterns.

Mature ovarian cystic teratomas are common benign ovarian neoplasm derived from germ cells. With increasing availability of ultrasound services even in developing countries, the diagnosis of benign ovarian tumour is made earlier and the size of the ovarian tumour at diagnosis is relatively small. It is unusual to find an ovarian cystic teratoma larger than 10 cm. We report a huge mature ovarian cystic teratoma in a multipara with a history of Hansen’s disease. We conclude that, in circumstances where women have restricted access to health care, the unusual finding of mature ovarian cystic teratoma larger than 10 cm is possible due to delayed presentation for diagnosis and treatment.

Our studies were conducted to explore the role of hepatic fatty acid-binding protein (L-FABP) in fatty acid transport to the nucleus. Purified rat L-FABP facilitated the specific interaction of [3H]oleic acid with the nuclei. L-FABP complexed with unlabeled oleic acid decreased the nuclear association of [3H]oleic acid:L-FABP; however, oleic acid-saturated bovine serum albumin (BSA) or fatty acid-free L-FABP did not. The peroxisome-proliferating agents LY171883, bezafibrate, and WY-14,643 were also effective competitors when complexed to L-FABP. Nuclease treatment did not affect the nuclear association of [3H]oleic acid:L-FABP; however, proteinase treatment of the nuclei abolished the binding. Nuclei incubated with fluorescein-conjugated L-FABP in the presence of oleic acid were highly fluorescent whereas no fluorescence was observed in reactions lacking oleic acid, suggesting that L-FABP itself was binding to the nuclei. The nuclear binding of FABP was concentration dependent, saturable, and competitive. LY189585, a ligand for L-FABP, also facilitated the nuclear binding of fluorescein-conjugated L-FABP, although it was less potent than oleic acid. A structural analog that does not bind L-FABP, LY163443, was relatively inactive in stimulating the nuclear binding. Potential interactions between L-FABP and nuclear proteins were analyzed by Far-Western blotting and identified a 33-kDa protein in the 500 mm NaCl extract of rat hepatocyte nuclei that bound strongly to biotinylated L-FABP. Oleic acid enhanced the interaction of L-FABP with the 33-kDa protein as well as other nuclear proteins. We propose that L-FABP is involved in communicating the state of fatty acid metabolism from the cytosol to the nucleus through an interaction with lipid mediators that are involved in nuclear signal transduction. —Lawrence, J. W., D. J. Kroll, and P. I. Eacho. Ligand-dependent interaction of hepatic fatty acid-binding protein with the nucleus. J. Lipid Res. 2000. 41: 1390–1401.

Mesenchymal stem cell (MSC)-based therapies have been proposed as novel treatments for intervertebral disc (IVD) degeneration. We have previously demonstrated that when MSCs are co-cultured with nucleus pulposus (NP) cells with direct cell-cell contact, they differentiate along the NP lineage and simultaneously stimulate the degenerate NP cell population to regain a normal (non-degenerate) phenotype, an effect which requires cell-cell communication. However, the mechanisms by which NP cells and MSCs interact in this system are currently unclear. Thus, in this study we investigated a range of potential mechanisms for exchange of cellular components or information that may direct these changes, including cell fusion, gap-junctional communication and exchange of membrane components by direct transfer or via microvesicle formation. Flow cytometry of fluorescently labeled MSCs and NP cells revealed evidence of some cell fusion and formation of gapjunctions, although at the three timepoints studied these phenomena were detectable only in a small proportion of cells. While these mechanisms may play a role in cell-cell communication, the data suggests they are not the predominant mechanism of interaction. However, flow cytometry of fluorescently dual-labeled cells showed that extensive bi-directional transfer of membrane components is operational during direct co-culture of MSCs and NP cells. Furthermore, there was also evidence for secretion and internalization of membrane-bound microvesicles by both cell types. Thus, this study highlights bi-directional intercellular transfer of membrane components as a possible mechanism of cellular communication between MSC and NP cells.

Factors that influence learning and the spread of behavior in wild animal populations are important for understanding species responses to changing environments and for species conservation. In populations of wildlife species that come into conflict with humans by raiding cultivated crops, simple models of exposure of individual animals to crops do not entirely explain the prevalence of crop raiding behavior. We investigated the influence of life history milestones using age and association patterns on the probability of being a crop raider among wild free ranging male African elephants; we focused on males because female elephants are not known to raid crops in our study population. We examined several features of an elephant association network; network density, community structure and association based on age similarity since they are known to influence the spread of behaviors in a population. We found that older males were more likely to be raiders than younger males, that males were more likely to be raiders when their closest associates were also raiders, and that males were more likely to be raiders when their second closest associates were raiders older than them. The male association network had sparse associations, a tendency for individuals similar in age and raiding status to associate, and a strong community structure. However, raiders were randomly distributed between communities. These features of the elephant association network may limit the spread of raiding behavior and likely determine the prevalence of raiding behavior in elephant populations. Our results suggest that social learning has a major influence on the acquisition of raiding behavior in younger males whereas life history factors are important drivers of raiding behavior in older males. Further, both life-history and network patterns may influence the acquisition and spread of complex behaviors in animal populations and provide insight on managing human-wildlife conflict.

Genetics plays an increasing role in modern life as evidenced by the development of revolutionary techniques such as CRISPR-based genome editing and the rise of personalized genome services. However, genetics is difficult to learn; known issues include its abstract nature, different scales, and technical language. Pedigree analysis is a convergence of these concepts, requiring use of multiple symbolic scales and understanding the relationships and nature of alleles, genes, and chromosomes. To measure student understanding of these concepts, as well as support biology educational reform toward student-centered instruction, we developed a formative assessment to provide reliable and valid evidence of student understanding, learning, and misconceptions for pedigree analysis. Nine multiple choice items targeted to four learning objectives were developed in an iterative process with faculty and student input. We designed distractor answers to capture common student misconceptions and deployed a novel statistical technique to assess the congruence of distractor language with targeted misconceptions. Psychometric analysis showed the instrument provides valid and reliable data and has utility to measure normalized learning gains. Finally, we employed cross-tabulation and distractor progression to identify several stable misconceptions that can be targeted for instructional intervention.

Outbreaks of insects and diseases are part of the natural disturbance regime of all forests. However, introduced pathogens have had outsized impacts on many dominant forest tree species over the past century. Mitigating these impacts and restoring these species are dilemmas of the modern era. Here, we review the ecological and economic impact of introduced pathogens, focusing on examples in North America. We then synthesize the successes and challenges of past biotechnological approaches and discuss the integration of genomics and biotechnology to help mitigate the effects of past and future pathogen invasions. These questions are considered in the context of the transgenic American chestnut, which is the most comprehensive example to date of how biotechnological tools have been used to address the impacts of introduced pathogens on naïve forest ecosystems. [ABSTRACT FROM AUTHOR]

Sustainability of the environment is one of the major problems facing African people, most especially Nigerians. It is unfortunate that women, by the nature of their daily activities of managing the homes and families are in touch with nature and environment and are at greater risk of health hazards and foetal damage. This paper focuses on the type of education that women need to instill in them knowledge and skills for handling environmental problems. The paper concludes with some recommendations that environmental courses, clubs, training and retraining of teachers, networking with NGOs on environmental education for sustainability should be implemented. To this end, the government should put in place laws and policies for effectiveness of environmental programmes.

The study aimed at examining the efficacy of Information and Communication Technology (ICT) in enhancing learning outcomes of students with hearing impairment in Ibadan. The study adopted a pretest, post-test, control group quasi-experimental research design. Purposive sampling techniques was used for the selection of thirty participants comprising of (15 males and 15 females) used for the study. The instrument used in carrying out the study was Economics Achievement Test (EAT) with reliability coefficient of 0.72. Analysis of covariance (ANCOVA) was used to analyse the null hypothesis generated and tested at 0.05 level of significance. The result showed that there is a significant main effect of ICT on the participants' learning outcomes in economics. Based on the findings of the study, it was recommended among others that teachers should utilize ICT during teaching learning process in the classroom and also go for in-service training to update their knowledge on the best way of handling ICT facilities to impact knowledge.

Predation can have cascading, regulatory effects across ecological communities. Knowledge of the diet of predators can therefore provide important information regarding their ecology and conservation, as well as their impacts on prey populations. Using scats collected during 2019–2023 and estimates of prey abundance from aerial surveys, we characterized prey consumption and preferences of the Vulnerable African lion Panthera leo population in Tsavo, Kenya. Biomass models applied to prey frequencies in scats revealed that > 85% of lion diet comprised large ungulates weighing > 150 kg. The Critically Endangered hirola Beatragus hunteri and Endangered Grevy's zebra Equus grevyi (species that were introduced in Tsavo as part of ex situ conservation programmes in the 1960s) were amongst the seven prey species, of 16 detected, that were preferred by lions. Our results potentially indicate a disproportionate impact of lion predation on the small hirola and Grevy's zebra populations. Preferential predation, coupled with high availability of alternative prey, may trap the small populations of hirola and Grevy's zebra within a predator pit. Our findings provide a better understanding of lion diet, optimal foraging and the potential effects predators can have on threatened and rare prey species in an important conservation landscape. Based on our findings, we recommend an observational study of the predation ecology of lions and other predators in this system, to provide information on age- and sex-specific predation rates on hirola and Grevy's zebra for a population viability analysis, to support the management of these two threatened and rare herbivores in Tsavo.