Plasminogen activator and collagenase production by cultured capillary endothelial cells

Cultured bovine capillary endothelial (BCE) cells produce low levels of collagenolytic activity and significant amounts of the serine protease plasminogen activator (PA). When grown in the presence of nanomolar quantities of the tumor promoter 12-O-tetradecanoyl phorbol-13-acetate (TPA), BCE cells produced 5-15 times more collagenolytic activity and 2-10 times more PA than untreated cells. The enhanced production of these enzymes was dependent on the dose of TPA used, with maximal response at 10(-7) to 10(-8) M. Phorbol didecanoate (PDD), an analog of TPA which is an active tumor promoter, also increased protease production. 4-O-methyl-TPA and 4α-PDD, two analogs of TPA which are inactive as tumor promoters, had no effect on protease production. Increased PA and collagenase activities were detected within 7.5 and 19 h, respectively, after the addition of TPA. The TPA-stimulated BCE cells synthesized a urokinase-type PA and a typical vertebrate collagenase. BCE cells were compared with bovine aortic endothelial (BAE) cells and bovine embryonic skin (BES) fibroblasts with respect to their production of protease in response to TPA. Under normal growth conditions, low levels of collagenolyic activity were detected in the culture fluids from BCE, BAE, and BES cells. BCE cells produced 5-13 times the basal levels of collagenolytic activity in response to TPA, whereas BAE cells and BES fibroblasts showed a minimal response to TPA. Both BCE and BAE cells exhibited relatively high basal levels of PA, the production of which was stimulated approximately threefold by the addition of TPA. The observation that BCE cells and not BAE cells produced high levels of both PA and collagenase activities in response to TPA demonstrates a significant difference between these two types of endothelial cells and suggests that the enhanced detectable activities are a property unique to bovine capillary and microvessel and endothelial cells.     

Novel Pancreatic Endocrine Maturation Pathways Identified by Genomic Profiling and Causal Reasoning

We have used a previously unavailable model of pancreatic development, derived in vitro from human embryonic stem cells, to capture a time-course of gene, miRNA and histone modification levels in pancreatic endocrine cells. We investigated whether it is possible to better understand, and hence control, the biological pathways leading to pancreatic endocrine formation by analysing this information and combining it with the available scientific literature to generate models using a casual reasoning approach. We show that the embryonic stem cell differentiation protocol is highly reproducible in producing endocrine precursor cells and generates cells that recapitulate many aspects of human embryonic pancreas development, including maturation into functional endocrine cells when transplanted into recipient animals. The availability of whole genome gene and miRNA expression data from the early stages of human pancreatic development will be of great benefit to those in the fields of developmental biology and diabetes research. Our causal reasoning algorithm suggested the involvement of novel gene networks, such as NEUROG3/E2F1/KDM5B and SOCS3/STAT3/IL-6, in endocrine cell development We experimentally investigated the role of the top-ranked prediction by showing that addition of exogenous IL-6 could affect the expression of the endocrine progenitor genes NEUROG3 and NKX2.2.     

Influence of the mother's reproductive state on the hormonal status of daughters in marmosets (Callithrix kuhlii)

Behavioral and endocrine suppression of reproduction in subordinate females produces the high reproductive skew that characterizes callitrichid primate mating systems. Snowdon et al. [American Journal of Primatology 31:11-21, 1993] reported that the eldest daughters in tamarin families exhibit further endocrinological suppression immediately following the birth of siblings, and suggested that dominant females exert greater control over subordinate endocrinology during this energetically challenging phase of reproduction. We monitored the endocrine status of five Wied's black tufted-ear marmoset daughters before and after their mother delivered infants by measuring concentrations of urinary estradiol (E(2)), pregnanediol glucuronide (PdG), testosterone (T), and cortisol (CORT). Samples were collected from marmoset daughters 4 weeks prior to and 9 weeks following three consecutive sibling-litter births when the daughters were prepubertal (M=6.1 months of age), peripubertal (M=11.9 months), and postpubertal (M=17.6 months). The birth of infants was associated with reduced ovarian steroid excretion only in the prepubertal daughters. In contrast, ovarian steroid levels tended to increase in the postpubertal daughters. Urinary E(2) and T levels in the postpubertal daughters were 73.8% and 37.6% higher, respectively, in the 3 weeks following the birth of infants, relative to prepartum levels. In addition, peak urinary PdG concentrations in peri- and postpubertal daughters were equivalent to luteal phase concentrations in nonpregnant, breeding adult females, and all of the peri- and postpubertal daughters showed clear ovulatory cycles. Cortisol excretion did not change in response to the reproductive status of the mother, nor did the concentrations change across age. Our data suggest that marmoset daughters of potential breeding age are not hormonally suppressed during the mother's peripartum period or her return to fertility. These findings provide an additional example of species diversity in the social regulation of reproduction in callitrichid primates.     

Treatment options after failure of radiation therapy-a review

Radioresistant or recurrent prostate cancer represents a serious health risk for approximately 20%-30% of patients treated with primary radiation therapy for clinically localized prostate cancer. The majority of patients exhibit large volume and poorly differentiated disease at the time of diagnosis, which limits the ability of salvage therapy to eradicate the cancer. Early detection with serum PSA monitoring and prostate needle biopsy following primary radiation therapy may identify residual adenocarcinoma at an earlier stage and increase the likelihood of successful salvage therapy. Radical prostatectomy, prostate cryoablation, and brachytherapy comprise the options for salvage treatment available for radiorecurrent prostate cancer. The goal of disease eradication must be balanced against the potential for serious treatment-related side effects. As a result, many patients receive noncurative therapy with androgen ablation despite the real risk of disease progression and mortality.     

Treatment with CRH-1 antagonist antalarmin reduces behavioral and endocrine responses to social stressors in marmosets (Callithrix kuhlii)

Corticotropin-releasing hormone (CRH) has multiple roles in coordinating the behavioral and endocrine responses to a host of environmental challenges, including social stressors. In the present study we evaluated the role of CRH in mediating responses to a moderate social stressor in Wied's black tufted-eared marmosets (Callithrix kuhlii). Male and female marmosets (n=14) were administered antalarmin (a selective CRH-1 receptor antagonist; 50 microg/kg, p.o.) or vehicle in a blind, counterbalanced, crossover design. One hr after treatment, marmosets were separated from long-term pairmates and then housed alone in a novel enclosure for 7 hr. Behavior was recorded during separation and upon reunion with the partner, and urine samples for cortisol assay collected before, during, and after the intervention. Separation from partners elevated urinary cortisol concentrations over baseline for both conditions, but antalarmin treatment reduced the magnitude of the elevation. Antalarmin also lowered rates of behavioral patterns associated with arousal (alarm and "e-e" vocalizations, object manipulate/chew), but had no effect on contact calls, locomotory activity or alertness. Although most patterns of social behavior upon reunion with the partner were not affected by antalarmin, antalarmin-treated marmosets displayed more sexual behavior (mounts and copulations) upon reunion. These data indicate that antagonism of the CRH-1 receptor acts to reduce the magnitude of both endocrine and behavioral responses to a moderate social stressor without causing any overall reduction in alertness or general activity. This supports the hypothesis that CRH, acting through its type 1 receptor, is involved in coordinating the responses to anxiety-producing events. These results further suggest that the marmoset is a useful model for exploration of the role of CRH in mediating the behavioral and neuroendocrine responses to psychosocial stressors, particularly in the context of heterosexual social relationships.     

Nighttime Wakefulness Associated with Infant Rearing in Callithrix kuhlii

Parent-infant cosleeping occurs in human and nonhuman primates, yet studies on the impact of cosleeping on parental sleep patterns have been limited to human mothers. We examined the effects of cosleeping on the nighttime wakefulness of a biparental New World primate, Wied's black tufted-ear marmoset (Callithrix kuhlii). We compared the sleep patterns of marmoset parents caring for young infants to those without infants, using an 8 mm videocamera and timelapse VCR under infrared illumination. The presence of young infants significantly impacted the sleep of mothers but not fathers. In fact, mothers rearing young infants were awake >3 times as often as mothers without infants. We also examined the nighttime wakefulness of marmoset parents across the first 9 weeks of infant life (birth through weaning). Although callitrichid mothers tend to reduce their daytime investment in offspring very early in infant life by relinquishing the care of infants to fathers and alloparents, increased nighttime wakefulness was not limited to the early postpartum period for the mothers. Instead, mothers exhibited more nighttime wakefulness than fathers did across the first 9 weeks of infant life. Our results indicate that the presence of infants has a greater impact on the sleep patterns of Callithrix kuhlii mothers than fathers, suggesting that mothers are more involved in infant care than previously realized and that fathers are not nearly as involved in nighttime care as their behavior during the day would suggest.     

A common polygenic basis for quinine and PROP avoidance in mice

Inbred strains of mice (Mus musculus) differ greatly in ability to taste various bitter compounds. For some compounds, the differences result from allelic variation at a single locus. However, segregation patterns incompatible with monogenic inheritance have been found for quinine avoidance. The Soa bitter sensitivity locus exerts some influence on this phenotype, but an unknown number of other loci also contribute. Relative avoidance patterns for quinine sulfate in panels of naive inbred strains resembled avoidance patterns for 6-n-propyl-2- thiouracil (PROP), suggesting a common genetic basis. In particular, C57BL/6J mice strongly avoided both 0.1 mM quinine sulfate and 1 mM PROP in two-bottle preference tests, whereas C3H/HeJ mice were indifferent to both. Therefore, 12 BXH/Ty recombinant inbred strains, derived from these strains, were tested with both solutions to begin identification of the unknown bitter loci. Naive mice were tested for four consecutive days with each compound (order counterbalanced). Some BXH/Ty strain means resembled those of the parent strains, but others were intermediate. This indicated recombination among loci affecting avoidance, and therefore polygenic inheritance. The strain means were highly correlated across compounds (r = 0.98), suggesting that the same polygenes controlled both phenotypes. The BXH/Ty means for both compounds were then compared with the strain genotypes at 212 chromosome position markers distributed throughout the genome. Eight markers on five chromosomes (3, 6, 7, 8 and 9) yielded significant correlations. Six of the markers were correlated with both phenotypes, again suggesting common polygenic inheritance. The marker with the highest correlation was Prp, tightly linked to Soa on chromosome 6. The correlated marker regions likely contain quantitative trait loci affecting bitter avoidance. The phenotypic similarity of PROP to quinine, rather than to phenylthiourea, apparently stemming from a common polygenic basis, indicates a difference between mice and humans in gustatory organization related to bitters.