Actin filament dynamics impacts keratinocyte stem cell maintenance.

Cultured human epidermal keratinocyte stem cells (holoclones) are crucial for regenerative medicine for burns and genetic disorders. In serial culture, holoclones progressively lose their proliferative capacity to become transient amplifying cells with limited growth (paraclones), a phenomenon termed clonal conversion. Although it negatively impacts the culture lifespan and the success of cell transplantation, little is known on the molecular mechanism underlying clonal conversion. Here, we show that holoclones and paraclones differ in their actin filament organization, with actin bundles distributed radially in holoclones and circumferentially in paraclones. Moreover, actin organization sets the stage for a differing response to epidermal growth factor (EGF), since EGF signalling induces a rapid expansion of colony size in holoclones and a significant reduction in paraclones. Furthermore, inhibition of PI3K or Rac1 in holoclones results in the reorganization of actin filaments in a pattern that is similar to that of paraclones. Importantly, continuous Rac1 inhibition in holoclones results in clonal conversion and reduction of growth potential. Together, our data connect loss of stem cells to EGF-induced colony dynamics governed by Rac1.

Contrast ultrasound: a simple-to-use phase-shifting medium offers saline infusion sonography-like images.

OBJECTIVE: To test the ability of a novel phase-shifting medium (PSM) to provide sustained distension of the uterine cavity and produce saline infusion sonography (SIS)-like images in a simplified contrast ultrasound procedure. DESIGN: Prospective pilot feasibility trial of a new diagnostic procedure, contrast ultrasound. SETTING: Clinical reproductive endocrine and infertility unit of regional teaching hospital. PATIENT(S): Twenty-six asymptomatic infertile women (group I) and 27 women presenting with dysfunctional uterine bleeding (DUB) who were scheduled for exploratory surgery (group II). INTERVENTION(S): All women who were temporarily on oral contraceptive first had a regular pelvic ultrasound followed by the intrauterine instillation of up to 3 mL PSM, using a regular insemination catheter, after which all instruments were removed and a regular ultrasound was performed again. RESULT(S): In all 53 women, intrauterine instillation of 1-3 mL PSM resulted in a 3-7 mm uterine distension, sufficient to produce SIS-like images of the uterine cavity that lasted 7-10 min. Contrast ultrasound revealed an endometrial polyp in 3 asymptomatic women of group I. In group II. 12 of 14 women (86%) whose vaginal ultrasound were positive or dubious had positive findings with contrast ultrasound; 9 of 12 patients whose vaginal ultrasounds were negative also had positive contrast ultrasound findings. All the positive and negative findings of contrast ultrasound made in group II were confirmed anatomically (sensitivity and specificity of 100%), whereas the correlation for standard vaginal ultrasound was markedly lower at 57.1% and 85.7%, respectively. Most patients (46 of 53) reported no discomfort during or after the procedure, and 7 women described the procedure as mildly uncomfortable. CONCLUSION(S): Contrast ultrasound, a novel simple diagnostic procedure conducted after intrauterine instillation of 1-3 mL PSM using a simple plastic catheter, delivered SIS-quality images in asymptomatic (group I) and symptomatic (group II) patients while retaining the simplicity of standard ultrasound. We therefore foresee broad application of contrast ultrasound for sensitive and specific assessment for uterine pathologies in the physician's office.

Ultrasound versus biological markers in the evaluation of periportal fibrosis in human Schistosoma mansoni

In this paper, the authors review the literature and share their experience of the principal biological markers of fibrosis for the evaluation of periportal fibrosis (PPF) caused by mansoni schistosomiasis. These biological markers are compared to diagnostic ultrasound (US) scans as means of grading PPF. We also review procollagen type I and III, collagen type IV, laminin, hyaluronic acid (HA), immunoglobulin G, platelets, aspartate aminotransferase to platelet ratio index (APRI) and gamma-glutamyl transpeptidase as markers of the disease. Although there are several good markers for evaluating PPF and portal hypertension, such as HA, platelets or APRI, none can yet replace US. These markers may, however, be used to identify patients at greater risk of developing advanced disease in endemic areas and determine who will need further care and US studies.

Ultrasound biomicroscopy evaluation of anterior extension in retinoblastoma: a clinicopathological study.

BACKGROUND: Extension of retinoblastoma cells into the posterior chamber is a criterion for group E according to the international classification of intraocular retinoblastoma. Currently, the anterior extension of retinoblastoma is based on the presence of tumour cells in the anterior chamber assessed by biomicroscopy. AIM: To determine the value of ultrasound biomicroscopy (UBM) in the assessment of posterior chamber involvement in advanced retinoblastoma. METHODS: Retrospective review of all retinoblastoma cases enucleated at the Jules Gonin Eye Hospital from January 1996 to December 2009 for which UBM (35 MHz) evaluation was available. The patients' records were reviewed for patient and tumour features and histopathological findings. UBM findings were compared with histopathological features. RESULTS: UBM documentation was available in 31 cases. Retinoblastoma was detected by UBM in the posterior chamber in 18 cases and was absent in 13 cases while histopathological analysis demonstrated its presence in the posterior chamber in 22 cases and its absence in 9 cases. Among the 18 UBM-positive cases, 7 had biomicroscopic detectable involvement of the anterior chamber. There was a significant correlation between echodensities consistent with retinoblastoma on UBM in the posterior chamber and histopathological tumorous involvement of the posterior chamber (p=0.0001). The sensitivity of UBM in the assessment of posterior chamber invasion by retinoblastoma was 81% and the specificity was 100%. CONCLUSION: In selected cases of advanced retinoblastoma, UBM appears to represent a valuable tool in the precise evaluation of anterior extension of disease, with good sensitivity and specificity for the assessment of posterior chamber involvement. UBM may provide useful criteria governing the indication for enucleation.

Induction and maintenance of protective CD8+ T cells against malaria liver stages: implications for vaccine development

CD8+ T cells against malaria liver stages represent a major protective immune mechanism against infection. Following induction in the peripheral lymph nodes by dendritic cells (DCs), these CD8+ T cells migrate to the liver and eliminate parasite infected hepatocytes. The processing and presentation of sporozoite antigen requires TAP mediated transport of major histocompatibility complex class I epitopes to the endoplasmic reticulum. Importantly, in DCs this process is also dependent on endosome-mediated cross presentation while this mechanism is not required for epitope presentation on hepatocytes. Protective CD8+ T cell responses are strongly dependent on the presence of CD4+ T cells and the capacity of sporozoite antigen to persist for a prolonged period of time. While human trials with subunit vaccines capable of inducing antibodies and CD4+ T cell responses have yielded encouraging results, an effective anti-malaria vaccine will likely require vaccine constructs designed to induce protective CD8+ T cells against malaria liver stages.

Performance de l'estimation du poids foetal par echographie pour les foetus < or = 2000 g et les macrosomes (> or = 4000 g) [Performance of ultrasound estimation of fetal weight in fetuses weighing < or = 2000 g and more than 4000 g].

Decreasing perinatal morbidity and mortality is one of the main goals of obstetrics. Prognosis of preterm births depends on gestational age and birthweight. Multidisciplinary management is discussed with the parents according to these two parameters. In other circumstances, a suspected macrosomy will influence the management of the last weeks of pregnancy. Induction of labor or Cesarean delivery will be considered to avoid shoulder dystocia, brachial plexus injury or perinatal asphyxia. Birthweight needs to be estimated with accuracy, and this article describes the efficiency of various ultrasound weight estimation formulae for small and large fetuses.

Quantitative ultrasound of bone in institutionalized elderly women: a cross-sectional and longitudinal study.

Quantitative ultrasound of bone is a promising method for bone assessment: radiation-free, portable and predictive of hip fracture. Its portability allowed us to study the relationships between ultrasonic parameters of bone with age and with non-vertebral fractures in elderly women living in 19 nursing homes. Broadband ultrasound attenuation (BUA) and speed of sound (SOS) of the calcaneus were measured (and the stiffness index calculated) in a sample of 270 institutionalized women, aged 85 +/- 7 years, using an Achilles bone densitometer (Lunar). The effects of age, history of non-vertebral and non-traumatic fractures, body mass index, triceps skinfold and arm circumference were assessed on BUA, SOS and stiffness index. Furthermore, to evaluate longitudinally the influence of aging on the ultrasound parameters of bone, 60 subjects from the same group had a second ultrasound measurement after 1 year. The cross-sectional analysis of the data on all 270 women showed a significant decrease (p < 0.001) with age in BUA, SOS and stiffness index (-0.47%, -0.06%, and -1.01% respectively per year). In the 94 women, (35%) with a history of previous non-vertebral fractures, ultrasound parameters were significantly lower (p < 0.0001) than in the 176 women with no history of fracture (-8.3% for BUA, -1.3% for SOS, -18.9% for stiffness index). In contrast, there was no significant difference in anthropometric measurements between the groups with and without previous non-vertebral fractures, although the measurements decreased significantly with age. In the longitudinal study, repeated quantitative ultrasound after 11.4 +/- 0.8 months showed no significant decrease in BUA (-1%) but a significant decrease in SOS (-0.3%, p < 0.0001) and in stiffness index (-3.6%, p < 0.0002). In conclusion, quantitative ultrasound of the calcaneus measures properties of bone which continue to decline in institutionalized elderly women, and is able to discriminate women with previous non-vertebral fractures.

Osteoprotegerin and bone mineral density in hemodiafiltration patients.

A newly identified cytokine, osteoprotegerin (OPG) appears to be involved in the regulation of bone remodeling. In vitro studies suggest that OPG, a soluble member of the TNF receptor family of proteins, inhibits osteoclastogenesis by interrupting the intercellular signaling between osteoblastic stromal cells and osteoclast progenitors. As patients with chronic renal failure (CRF) often have renal osteodystrophy (ROD), we investigated the role of osteoprotegerin (OPG) in ROD, and investigated whether there was any relationship between serum OPG, intact parathyroid (PTH) (iPTH), vitamin D, and trabecular bone. Serum OPG combined with iPTH might be a useful tool in the noninvasive diagnosis of ROD, at least in cases in which the range of PTH values compromises reliable diagnosis. Thirty-six patients on maintenance hemodiafiltration (HDF) and a control group of 36 age and sex matched healthy subjects with no known metabolic bone disease were studied. The following assays were made on serum: iPTH, osteocalcin (BGP), bone alkaline phosphatase, 25(OH)-cholecalciferol, calcium, phosphate, OPG, IGF-1, estradiol, and free testosterone. Serum Ca++, P, B-ALP, BGP, IGF-1, iPTH, and OPG levels were significantly higher in HDF patients than in controls, while DXA measurements and quantitative ultrasound (QUS) parameters were significantly lower. On grouping patients according to their mean OPG levels, we observed significantly lower serum IGF-1, vitamin D3 concentrations, and lumbar spine and hip bone mineral density in the high OPG groups. No correlation was found between OPG and bone turnover markers, whereas a negative correlation was found between serum OPG and IGF-1 levels (r=-0.64, p=0.032). Serum iPTH concentrations were positively correlated with bone alkaline phosphatase (B-ALP) (r=0.69, p=0.038) and BGP (r=0.92, p<0.001). The findings made suggest that an increase in OPG levels may be a compensatory response to elevated bone loss. The low bone mineral density (BMD) levels found in the high OPG group might have been due to the significant decrease in serum IGF-1 and vitamin D3 observed. In conclusion, the findings made in the present study demonstrate that increased OPG in hemodiafiltration patients is only partly due to decreased renal clearance. As it may partly reflect a compensatory response to increased bone loss, this parameter might be helpful in the identification of patients with a marked reduction in trabecular BMD.

Mate choice evolution, dominance effects, and the maintenance of genetic variation.

Female mate choice influences the maintenance of genetic variation by altering the mating success of males with different genotypes. The evolution of preferences themselves, on the other hand, depends on genetic variation present in the population. Few models have tracked this feedback between a choice gene and its effects on genetic variation, in particular when genes that determine offspring viability and attractiveness have dominance effects. Here we build a population genetic model that allows comparing the evolution of various choice rules in a single framework. We first consider preferences for good genes and show that focused preferences for homozygotes evolve more easily than broad preferences, which allow heterozygous males high mating success too. This occurs despite better maintenance of genetic diversity in the latter scenario, and we discuss why empirical findings of superior mating success of heterozygous males consequently do not immediately lead to a better understanding of the lek paradox. Our results thus suggest that the mechanisms that help maintain genetic diversity also have a flipside of making female choice an inaccurate means of producing the desired kind of offspring. We then consider preferences for heterozygosity per se, and show that these evolve only under very special conditions. Choice for compatible genotypes can evolve but its selective advantage diminishes quickly due to frequency-dependent selection. Finally, we show that our model reproduces earlier results on selfing, when the female choice strategy produces assortative mating. Overall, our model indicates that various forms of heterozygote-favouring (or variable) female choice pose a problem for the theory of sexual ornamentation based on indirect benefits, rather than a solution.

Development and maintenance of the neuronal cytoskeleton in aggregated cell cultures of fetal rat telencephalon and influence of elevated K+ concentrations.

Serum-free aggregating cell cultures of fetal rat telencephalon were examined by biochemical and immunocytochemical methods for their development-dependent expression of several cytoskeletal proteins, including the heavy- and medium-sized neurofilament subunits (H-NF and M-NF, respectively); brain spectrin; synapsin I; beta-tubulin; and the microtubule-associated proteins (MAPs) 1, 2, and 5 and tau protein. It was found that with time in culture the levels of most of these cytoskeletal proteins increased greatly, with the exceptions of the particular beta-tubulin form studied, which remained unchanged, and MAP 5, which greatly decreased. Among the neurofilament proteins, expression of M-NF preceded that of H-NF, with the latter being detectable only after approximately 3 weeks in culture. Furthermore, MAP 2 and tau protein showed a development-dependent change in expression from the juvenile toward the adult form. The comparison of these developmental changes in cytoskeletal protein levels with those observed in rat brain tissue revealed that protein expression in aggregate cultures is nearly identical to that in vivo during maturation of the neuronal cytoskeleton. Aggregate cultures deprived of glial cells, i.e., neuron-enriched cultures prepared by treating early cultures with the antimitotic drug cytosine arabinoside, exhibited pronounced deficits in M-NF, H-NF, MAP 2, MAP 1, synapsin I, and brain spectrin, with increased levels of a 145-kDa brain spectrin breakdown product. These adverse effects of glial cell deprivation could be reversed by the maintenance of neuron-enriched cultures at elevated concentrations of KCl (30 mM). This chronic treatment had to be started at an early developmental stage to be effective, a finding suggesting that sustained depolarization by KCl is able to enhance the developmental expression and maturation of the neuronal cytoskeleton.

Dyssegmental dysplasia, Silverman-Handmaker type: prenatal ultrasound findings and molecular analysis.

OBJECTIVES: The objective of this study is to describe the prenatal sonographic features and the results of DNA analysis on three fetuses with dyssegmental dysplasia, Silverman-Handmaker type (DD-SH). METHODS: A retrospective review of three fetuses with confirmed DD-SH was conducted. The fetal ultrasound findings, the radiological characteristics, and the results of the mutation analysis of the heparan sulphate perlecan gene 2 (HSPG2) were reviewed. RESULTS: There were three cases in two families with DD-SH diagnosed prenatally. The main prenatal ultrasound and the radiological features of DD-SH were severe limb shortening and vertebral segmentation and fusion defects (anisospondyly). The DNA analysis of the HSPG2 gene showed that the two affected fetuses in a nonconsanguineous family had a compound heterozygote for the c.646G > T transversion in exon 7 and a c.5788C > T transition in exon 46. The fetus born to the consanguineous couple had a homozygous mutation c.1356-27_1507 + 59del. CONCLUSION: DD-SH can be diagnosed prenatally using fetal ultrasound as early as 13 weeks. Xrays and DNA analysis of the HSPG2 gene are important for the confirmation of the diagnosis and for the preimplantation and prenatal diagnosis in pregnancies at risk. © 2013 John Wiley & Sons, Ltd.

Intraovarian sperm storage in Helicolenus dactylopterus dactylopterus fertilitzation, crypt formation and maintenance of stored sperm

The females of the bluemouth rockfish, Helicolenus dactylopterus dactylopterus (DelaRoche, 1809), store sperm within their ovaries for periods of up to 10 months. Twenty six females with standard lengths between 152 and 257 mm and six males with standard lengths between 253 and 209 mm were caught storage crypts with stored spermatozoa and to describe their evolution over the year. After internal fertilization and once sperm reaches the ovary, a crypt forms probably by an epithelial inclusion at the base of the lamellae of one or several spermatozoa groups that are floating freely in the interlamellar space of the ovarian lumen. Stored spermatozoa have a large cytoplasm bag surrounding their heads. This bag could serve as a nutritive reservoir during the long storage period. Many desmosonal and tight junctions between the crypt cells ensure tha male sex cells are protected against the female immune system

Draft genome of the Leptospira interrogans strains, Acegua, RCA, Prea, and Capivara, obtained from wildlife maintenance hosts and infected domestic animals

In the present paper, we announce new draft genomes of four Leptospira interrogans strains named Acegua, RCA, Prea, and Capivara. These strains were isolated in the state of Rio Grande do Sul, Brazil, from cattle, dog, Brazilian guinea pig, and capybara, respectively.