Statins significantly repress rotavirus replication through downregulation of cholesterol synthesis.

Rotavirus is the most common cause of severe diarrhea among infants and young children and is responsible for more than 200,000 pediatric deaths per year. There is currently no pharmacological treatment for rotavirus infection in clinical activity. Although cholesterol synthesis has been proven to play a key role in the infections of multiple viruses, little is known about the relationship between cholesterol biosynthesis and rotavirus replication. The models of rotavirus infected two cell lines and a human small intestinal organoid were used. We investigated the effects of cholesterol biosynthesis, including inhibition, enhancement, and their combinations on rotavirus replication on these models. The knockdown of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) was built by small hairpin RNAs in Caco2 cells. In all these models, inhibition of cholesterol synthesis by statins or HMGCR knockdown had a significant inhibitory effect on rotavirus replication. The result was further confirmed by the other inhibitors: 6-fluoromevalonate, Zaragozic acid A and U18666A, in the cholesterol biosynthesis pathway. Conversely, enhancement of cholesterol production increased rotavirus replication, suggesting that cholesterol homeostasis is relevant for rotavirus replication. The effects of all these compounds toward rotavirus were further confirmed with a clinical rotavirus isolate. We concluded that rotavirus replication is dependent on cholesterol biosynthesis. To be specific, inhibition of cholesterol synthesis can downregulate rotavirus replication; on the contrary, rotavirus replication is upregulated. Statin treatment is potentially an effective novel clinical anti-rotavirus strategy.

Pseudorabies virus production using a serum-free medium in fixed-bed bioreactors with low cell inoculum density.

Fixed-bed bioreactors packed with macrocarriers show great potential to be used for vaccine process development and large-scale production due to distinguishing features of low shear force, high cell adhering surface area, and easy replacement of culture media in situ. As an initial step of utilizing this type of bioreactors for Pseudorabies virus production (PRV) by African green monkey kidney (Vero) cells, we developed a tube-fixed-bed bioreactor in the previous study, which represents a scale-down model for further process optimization. By using this scale-down model, here we evaluated impacts of two strategies (use of serum-free medium and low cell inoculum density) on PRV production, which have benefits of simplifying downstream process and reducing risk of contamination. We first compared Vero cell cultures with different media, bioreactors and inoculum densities, and conclude that cell growth with serum-free medium is comparable to that with serum-containing medium in tube-fixed-bed bioreactor, and low inoculum density supports cell growth only in this bioreactor. Next, we applied serum-free medium and low inoculum cell density for PRV production. By optimization of time of infection (TOI), multiplicity of infection (MOI) and the harvesting strategy, we obtained total amount of virus particles ~ 9 log10 TCID50 at 5 days post-infection (dpi) in the tube-fixed-bed bioreactor. This process was then scaled up by 25-fold to a Xcell 1-L fixed-bed bioreactor, which yields totally virus particles of 10.5 log10 TCID50, corresponding to ~ 3 × 105 doses of vaccine. The process studied in this work holds promise to be developed as a generic platform for the production of vaccines for animal and human health.

Modeling variation in the growth of wild and captive juvenile vervet monkeys in relation to diet and resource availability.

OBJECTIVES: To compare longitudinal weight gain in captive and wild juvenile vervet monkeys and conduct an empirical assessment of different mechanistic growth models. METHODS: Weights were collected from two groups of captive monkeys and two consecutive cohorts of wild monkeys until the end of the juvenile period (~800 days). The captive groups were each fed different diets, while the wild groups experienced different ecological conditions. Three different growth curve models were compared. RESULTS: By 800 days, the wild juveniles were lighter, with a slower maximum growth rate, and reached asymptote earlier than their captive counterparts. There were overall differences in weight and growth rate across the two wild cohorts. This corresponded to differences in resource availability. There was considerable overlap in growth rate and predicted adult weight of male and females in the first, but not the second, wild cohort. Maternal parity was not influential. While the von Bertalanffy curve provided the best fit to the data sets modeled together, the Logistic curve best described growth in the wild cohorts when considered separately. CONCLUSIONS: The growth curves of the two captive cohorts are likely to lie near the maximum attainable by juvenile vervets. It may be helpful to include deviations from these rates when assessing the performance of wild vervet monkeys. The comparison of wild and captive juveniles confirmed the value of comparing different growth curve models, and an appreciation that the best models may well differ for different populations. Choice of mechanistic growth model can, therefore, be empirically justified, rather than theoretically predetermined.

Obesity and obesogenic growth are both highly heritable and modified by diet in a nonhuman primate model, the African green monkey (Chlorocebus aethiops sabaeus).

OBJECTIVE: In humans, the ontogeny of obesity throughout the life course and the genetics underlying it has been historically difficult to study. We compared, in a non-human primate model, the lifelong growth trajectories of obese and non-obese adults to assess the heritability of and map potential genomic regions implicated in growth and obesity. STUDY POPULATION: A total of 905 African green monkeys, or vervets (Chlorocebus aethiops sabaeus) (472 females, 433 males) from a pedigreed captive colony. METHODS: We measured fasted body weight (BW), crown-to-rump length (CRL), body-mass index (BMI) and waist circumference (WC) from 2000 to 2015. We used a longitudinal clustering algorithm to detect obesogenic growth, and logistic growth curves implemented in nonlinear mixed effects models to estimate three growth parameters. We used maximum likelihood variance decomposition methods to estimate the genetic contributions to obesity-related traits and growth parameters, including a test for the effects of a calorie-restricted dietary intervention. We used multipoint linkage analysis to map implicated genomic regions. RESULTS: All measurements were significantly influenced by sex, and with the exception of WC, also influenced by maternal and post-natal diet. Chronic obesity outcomes were significantly associated with a pattern of extended growth duration with slow growth rates for BW. After accounting for environmental influences, all measurements were found to have a significant genetic component to variability. Linkage analysis revealed several regions suggested to be linked to obesity-related traits that are also implicated in human obesity and metabolic disorders. CONCLUSIONS: As in humans, growth patterns in vervets have a significant impact on adult obesity and are largely under genetic control with some evidence for maternal and dietary programming. These results largely mirror findings from human research, but reflect shorter developmental periods, suggesting that the vervet offers a strong genetic model for elucidating the ontogeny of human obesity.

Genetic variation and gene expression across multiple tissues and developmental stages in a nonhuman primate.

By analyzing multitissue gene expression and genome-wide genetic variation data in samples from a vervet monkey pedigree, we generated a transcriptome resource and produced the first catalog of expression quantitative trait loci (eQTLs) in a nonhuman primate model. This catalog contains more genome-wide significant eQTLs per sample than comparable human resources and identifies sex- and age-related expression patterns. Findings include a master regulatory locus that likely has a role in immune function and a locus regulating hippocampal long noncoding RNAs (lncRNAs), whose expression correlates with hippocampal volume. This resource will facilitate genetic investigation of quantitative traits, including brain and behavioral phenotypes relevant to neuropsychiatric disorders.

A comparison of adult body size between captive and wild vervet monkeys (Chlorocebus aethiops sabaeus) on the island of St. Kitts.

Weight and 34 morphological measurements were obtained from 103 vervet monkeys living either in the wild or in captive colonies derived from the wild populations on the island of St. Kitts in the Eastern Caribbean. All measures were taken during the same week, eliminating bias that might result from changing seasonal environmental conditions. Vervets on St. Kitts are all descended from a small number of individuals brought to the island approximately 400 years ago from West Africa, thus eliminating bias that might result from subspecific size differences. We conducted a principal components analysis (PCA) and compared individual traits between captive and wild adult animals. Morphological measures such as body, arm, and leg length did not differ significantly between animals living in the wild and animals in captivity. Weight and measures indicating condition-including body mass index (BMI), chest, thigh, and upper arm girth were all higher for animals living in captivity. More consistent available food is probably the cause of differences in measures reflecting condition.

Alterations in levels and ratios of n-3 and n-6 polyunsaturated fatty acids in the temporal cortex and liver of vervet monkeys from birth to early adulthood.

Deficiencies in omega-3 (n-3) long chain polyunsaturated fatty acids (LC-PUFAs) and increases in the ratio of omega-6 (n-6) to n-3 LC-PUFAs in brain tissues and blood components have been associated with psychiatric and developmental disorders. Most studies have focused on n-3 LC-PUFA accumulation in the brain from birth until 2years of age, well before the symptomatic onset of such disorders. The current study addresses changes that occur in childhood and adolescence. Postmortem brain (cortical gray matter, inferior temporal lobe; n=50) and liver (n=60) from vervet monkeys fed a uniform diet from birth through young adulthood were collected from archived tissues. Lipids were extracted and fatty acid levels determined. There was a marked reduction in the ratio of n-6 LC-PUFAs, arachidonic acid (ARA) and adrenic acid (ADR), relative to the n-3 LC-PUFA, docosahexaenoic acid (DHA), in temporal cortex lipids from birth to puberty and then a more gradual decrease though adulthood. This decrease in ratio resulted from a 3-fold accumulation of DHA levels while concentrations of ARA remained constant. Early childhood through adolescence appears to be a critical period for DHA accretion in the cortex of vervet monkeys and may represent a vulnerable stage where lack of dietary n-3 LC-PUFAs impacts development in humans.

Canonical wnt signaling is required for commissural axon guidance.

Morphogens have been identified as guidance cues for postcrossing commissural axons in the spinal cord. Shh has a dual effect on postcrossing commissural axons: a direct repellent effect mediated by Hhip as a receptor, and an indirect effect by shaping a Wnt activity gradient. Wnts were shown to be attractants for postcrossing commissural axons in both chicken and mouse embryos. In mouse, the effects of Wnts on axon guidance were concluded to depend on the planar cell polarity (PCP) pathway. Canonical Wnt signaling was excluded based on the absence of axon guidance defects in mice lacking Lrp6 which is an obligatory coreceptor for Fzd in canonical Wnt signaling. In the loss-of-function studies reported here, we confirmed a role for the PCP pathway in postcrossing commissural axon guidance also in the chicken embryo. However, taking advantage of the precise temporal control of gene silencing provided by in ovo RNAi, we demonstrate that canonical Wnt signaling is also required for proper guidance of postcrossing commissural axons in the developing spinal cord. Thus, axon guidance does not seem to depend on any one of the classical Wnt signaling pathways but rather involve a network of Wnt receptors and downstream components.

Endocrine mechanisms of primate life history trade-offs: growth and reproductive maturation in vervet monkeys.

Life history theory predicts that the timing of maturation will result from a trade-off between growth and the age of first reproduction. This trade-off and its mechanisms of action are still poorly understood in many species and have not been well studied at the individual level. This study examined hypothesized trade-offs between growth and reproductive maturation in wild populations of vervet monkeys (Cercopithecus aethiops) from Kenya, East Africa. Individuals were sampled from four populations in widely separated sites differing in temperature, altitude, and rainfall. Biological samples and morphometric measures were collected from 50 adult males, 83 adult females, and 225 juveniles. Gonadal steroids and leptin levels were analyzed by radioimmunoassay of sera from 136 juvenile males and 90 juvenile females. Cross-sectional profiles of morphometric and endocrine data were used to assess the onset and cessation of growth in relation to sexual maturation. Gonadal steroids were used to assess sexual maturation and breeding onset. Leptin was used as an index of nutritional state. Estimates of mortality were derived from population age-structure. Across populations, higher resource productivity and nutrient status were associated with more rapid growth. Shorter growth duration was associated with earlier reproductive onset. These findings provide support for models of trade-offs between the timing of growth completion and reproductive onset, but they are contradicted by the evidence that reproduction precedes the cessation of growth in these populations. The biphasic actions of estradiol provide an alternative model and mechanism for the growth-reproduction trade-off.

Microarray analysis of developmental plasticity in monkey primary visual cortex.

We performed microarray gene expression analyses on the visual cortex of Old-World monkeys (Cercopithicus aethiops) in an effort to identify transcripts associated with developmental maturation and activity-driven changes during the visual critical period. Samples derived from normal animals and those subjected to monocular enucleation (ME) were hybridized to human Affymetrix HG-U95Av2 oligonucleotide microarrays (N = 12) and the results were independently validated by real-time quantitative RT-PCR. To identify genes exhibiting significant expression differences among our samples, the microarray hybridization data were processed with two software packages that use different analytical models (Affymetrix MicroArray Suite 5.0, dChip 1.2). We identified 108 transcripts within diverse functional categories that differed in their visual cortical expression at the height of the critical period when compared to adults. The expression levels of four transcripts were also globally modulated following ME during the critical period. These transcripts are particularly sensitive to ME during the critical period but are not significantly modulated in ME adults. Three of the ME-driven genes (NGFI-B, egr3, NARP) are known immediate-early genes (IEG) while the other (DUSP6) is a phosphatase that can regulate IEG expression. The putative biological significance of the ME-driven and developmentally regulated genes is discussed with respect to the critical period for activity-dependent visual cortical neuroplasticity.

Sensory regulation of immediate-early genes c-fos and zif268 in monkey visual cortex at birth and throughout the critical period.

The postnatal development of ocular dominance columns (ODCs) in monkey visual cortex provides an exquisite model for studying mechanisms of experience-guided neuronal plasticity. While the presence of columns at birth in Old World monkeys is now well established, it remains unclear whether cortical neurons at this early stage are capable of modulating gene expression in response to changing sensory conditions. Using a set of monocular deprivation and stimulation protocols, we examined activity-driven expression of the immediate-early genes (IEGs) c-fos and zif268 during the critical period of development. We observed well-delineated patterns of ODCs produced by sensory regulation of both IEGs throughout the critical period, starting as early as the first postnatal day. The expression levels are similar in layers II/II, IVC and VI throughout development, with no selective decline in the thalamorecepient layer (layer IVC) of adult monkeys. A narrow strip of non-columnar c-Fos expression was observed at the border of layers IVC and V. Our results show that neurons in monkey visual cortex are equipped at birth with the molecular machinery for coupling sensory inputs to active genomic responses and that this responsivity extends throughout the critical period. The findings are discussed within the context of a possible role for IEGs in sensory-driven cortical plasticity during development.

Age-related expression of the CD15 (3-fucosyl-N-acetyl-lactosamine) epitope in the monkey (Cercopithecus aethiops aethiops L.) lateral geniculate nucleus.

The profile of the CD15 epitope-expression was studied in the lateral geniculate nucleus (LGN) of a monkey species (Cercopithecus aethiops aethiops L.) during the peri- and postnatal development. Intense labeling of the neuropil and of astroglial cells was detected. Morphological examination of the spatio-temporal appearance of the CD15-expression showed different consecutive patterns of expression: expression of the CD15 epitope was high one week prenatally and during the first postnatal weeks. During this perinatal period it was mainly expressed in the interlaminar zone of the magnocellular portion of the LGN. This was followed by a time period when immunoreactivity gradually decreased to become almost absent by approximately 10 weeks of age. Toward adulthood a different pattern of immunoreactivity occurred, revealing a pattern of lamination that was attributable to CD15 positive astrocytes, most prominent in the cellular layers 1 and 2. They were later on seen spread out through the entire LGN such that the LGN of adult animals was entirely filled with positive material. From the temporal correlation of CD15-expression and LGN histogenesis it is concluded that the high level of CD15, that is observed in the neuropil during the perinatal period, matches with high synaptic plasticity within the visual system; in contrast, low levels of CD15-expression correlate with synaptic reorganization processes.

Parathyroid hormone, ionised calcium, and potentially interacting variables in plasma of an Old World primate.

Bone turnover and calcium homeostasis in man can only be modelled validly in Old World nonhuman primates. In order to interpret the models it is necessary to establish endocrine and biochemical parameters of bone mineral metabolism. This report is probably the first description of acute phase parathyroid responses to manipulations of blood ionised calcium, and of reference values for potentially interacting variables, in vervet monkeys. Plasma parathyroid hormone concentrations were measured in vervets under defined conditions, and ranges reported as normal for other nonhuman primates and man are summarised.

[Growth curves of body weight and their relationship to sexual maturity in laboratory-bred male African green monkeys (Cercopithecus aethiops)].

Nonlinear growth models having a three- or four-parameter family were applied to individual body weight data of 5 male African green monkeys for estimating their growth patterns. Body weight was measured from birth to six years of age and 58 to 114 data items per monkey were collected. The average body weight at birth was 360g with the standard deviation of +/- 25g, 4.54 +/- 0.29 kg at five years of age, and 4.50 +/- 0.12 kg at six years of age at which point body weight was judged to have reached a plateau. Five growth models (Gompertz, Logistic, Richards, Bertalanffy and Brody) were applied to the growth data in this study. As a result, two (Gompertz and Logistic) of the five models were found applicable to all data from the five monkeys. However, the coefficient of determination (R2) obtained by application of the two models were not so large (0.919 +/- 0.05 in Gompertz, 0.889 +/- 0.01 in Logistic). Therefore the data were divided into two groups according to monkey age: the first group being from monkeys between birth and 2 years 10 months of age and the second group was from monkeys older than 2 years 10 months of age. The Gompertz model fitted best the data of the first group in four of the five animals (R2 = 0.982 +/- 0.011). The age at the inflexion point in the Gompertz model nearly corresponded to the age of weaning. The Logistic model was most suitable for the date of the second group in all five animals (R2 = 0.955 +/- 0.038).(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of size on vervet (Cercopithecus aethiops) gait parameters: a longitudinal approach.

Changes in the values of certain locomotor parameters were analyzed over a range of speeds for five immature vervet monkeys sampled at 6 month intervals over approximately a 3 year period. Lateral and diagonal sequence walking gaits and transverse and rotary gallops were commonly used. The monkeys switched from walking to galloping at long cycle durations for their mass, although, as a group, their transition speeds were in agreement with data from other mammals. However, for individual monkeys, transition speed was not consistently dependent on body mass. Cycle and stance durations generally increased with increasing size at each speed for each animal, with the greatest increases occurring at slower speeds. Swing durations increased slightly with size. For any particular individual, speed was highly predictable from cycle (or stance) duration and body mass (or age). However, the multiple regression equations for each animal were significantly different from each other, suggesting that no single equation is satisfactory for all of the individuals within a species.

Effects of growth and speed on hindlimb joint angular displacement patterns in vervet monkeys (Cercopithecus aethiops).

Hip, knee, and ankle joint displacement patterns are compared across both age and speed for five immature vervet monkeys sampled approximately every 6 months over a 3 year period. The analysis indicated that, as a group, the animals displayed no consistent changes in joint patterns as they grew. However, individual animals showed consistent patterns. There were also no consistent effects of size across animals at the walk-gallop transition. This is contrary to McMahon's prediction (J. Appl. Physiol. 39:619-627, 1975) based upon his elastic-similarity model of animal scaling. With increasing speed, when symmetrical gaits were used, all of the animals tended to show a decrease in the relative positions of the hip, knee, and ankle maximum values. Furthermore, across the walk-gallop transition, the animals tended to show a decrease in the range of ankle and knee movements.

[Spheno-occipital synchondrosis--a fluorescence and polarization microscopy study in Cercopithecus aethiops monkeys]. / Die Synchondrosis sphenooccipitalis--eine fluoreszenz- und polarisationsmikroskopische Untersuchung am Cercopithecus-aethiops-Affen.

On 17 Cercopithecus aethiops monkeys we investigated with the method of the polychromic sequential dye marking system the histomorphology as well as the dynamics of growth and calcification of the spheno-occipital synchondrosis. The age of the animals ranged from change to teeth to late adolescence. The animals had been divided into four different groups: I: late change of teeth; II: young adult; III: fully grown; IV: late adolescence. In the first group the spheno-occipital synchondrosis showed no ossification in the gap which is filled with cartilage. It showed the characteristic structure with a central zone of equally distributed chondrocytes. Adjacent to this we found a zone of proliferation cell hypotrophy and cell degeneration. With increasing age there is decrease of the density of cells (groups II to IV) and after change to teeth the synchondrosis starts to ossify (group II). Due to the ossification the synchondrosis subdivides into different cartilage regions. We found that the closing of the synchondrosis started in the cranial region and progressed toward the caudal region. During this procedure the synchondrosis never ossified completely. Several cartilage regions persisted uncalcified until late adolescence. Interstitial growth of the synchondrosis was found until the end of the change of the teeth (group I). This growth which was always found in a sagittal direction ceased after bony connections had been formed between both poles of the synchondrosis. The sphenoidal and occipital pole of the synchondrosis showed equal growth potential.

[Growth curves of body weight changes in laboratory-bred female African green monkeys (Cercopithecus aethiops)].

Nonlinear growth models having three or four parameter family were applied to individual weight data of female African green monkeys for estimating their growth pattern. The body weight was measured continuously from birth to six years of age with five female laboratory-bred monkeys. A total of 95 weight data were collected from each monkey. The average body weight was 330 g with the standard deviation of +/- 15 g at birth, and 2.71 +/- 0.33 kg at four years of age. The body weight of female African green monkeys was judged to reach a plateau after about four years of age. Five growth models (Gompertz, Logistic, Richards, Bertalanffy, Brody) were applied to these weight to age data. The most suitable coefficient of determination between growth data and growth model was obtained by the application of Gompertz equation. Three parameters of Gompertz equation, mature size (A), rate of maturing (K) and inflexion point (e-1 A) were analyzed in relation to age of menarche. Strong correlations between age of menarche and maturing rate, as well as between age of menarche and inflexion point were observed.

Immunohistochemical study of ontogeny and phylogeny of a terminal N-acetylglucosamine cluster antigen.

In this work an immunohistochemical method was used to study the ontogeny and phylogeny of a terminal N-acetylglucosamine (GlcNAc) cluster antigen which is an epitope(s) of highly branched N-linked oligosaccharides terminating in GlcNAc residues. The ontogenic studies demonstrated that expression of the antigen is developmentally regulated in lymphocytes, epithelial cells of endodermal origin and kidney mesangial cells of the chicken. The antigen was found in several other avian species studied, namely, the Japanese quail, duck, goose and turkey. Furthermore, the distribution of the antigen in all these species was similar. In adult animals, it was found in bursal and thymic lymphocytes, macrophages, spleen reticulum cells, epithelial cells of the intestine and bronchioles and capillary endothelial cells. The antigen was also detected in epithelial cells of the gastrointestinal tract of several lower vertebrates studied: the amphibian (frog), reptile (chameleon) and fish (rainbow trout). It was undetectable in various organs of the human, African green monkey, calf, pig, rat and guinea-pig, but was found in the intestinal epithelial cells of ten mouse strains. It is likely that biosynthetic processing leading to the formation of highly branched N-linked glycans terminating in GlcNAc residues is conserved during evolution in birds and other lower vertebrates.