Two mechanistically distinct forms of endocytosis in adrenal chromaffin cells: Differential effects of SH3 domains and amphiphysin antagonism

We previously identified two forms of endocytosis using capacitance measurements in chromaffin cells: rapid endocytosis (RE), dynamin-1 dependent but clathrin-independent and slow endocytosis (SE), dynamin-2 and clathrin-dependent. Various recombinant SH3 domains that interact with the proline-rich domain of dynamin were introduced into single cells via the patch pipette. GST-SH3 domains of amphiphysin-1, intersectin-IC, and endophilin-I inhibited SE but had no effect on RE. Grb2-SH3 (N-terminal) or a mutant of amphiphysin-1-SH3 was inactive on either process. These data confirm that dynamin-1 dependent RE is independent of clathrin and show that amphiphysin is exclusively associated with clathrin and dynamin-2-dependent SE.     

Human embryonic stem cell-derived neural precursor transplants in collagen scaffolds promote recovery in injured rat spinal cord

Background aimsSeveral studies have reported functional improvement after transplantation of in vivo-derived neural progenitor cells (NPC) into injured spinal cord. However, the potential of human embryonic stem cell-derived NPC (hESC-NPC) as a tool for cell replacement of spinal cord injury (SCI) should be considered.MethodsWe report on the generation of NPC as neural-like tubes in adherent and feeder-free hESC using a defined media supplemented with growth factors, and their transplantation in collagen scaffolds in adult rats subjected to midline lateral hemisection SCI.ResultshESC-NPC were highly expressed molecular features of NPC such as Nestin, Sox1 and Pax6. Furthermore, these cells exhibited the multipotential characteristic of differentiating into neurons and glials in vitro. Implantation of xenografted hESC-NPC into the spinal cord with collagen scaffold improved the recovery of hindlimb locomotor function and sensory responses in an adult rat model of SCI. Analysis of transplanted cells showed migration toward the spinal cord and both neural and glial differentiation in vivo.ConclusionsThese findings show that transplantation of hESC-NPC in collagen scaffolds into an injured spinal cord may provide a new approach to SCI.     

Design of Localized Surface Plasmon Resonance (LSPR) Biosensor for Immunodiagnostic of E. coli O157:H7 Using Gold Nanoparticles Conjugated to the Chicken Antibody

E. coli O157:H7 is one of the most important pathogens in food-borne diseases and is the main cause of the pseudo pandemic development of hemorrhagic colitis and hemolytic uremic syndrome. Also E. coli O157:H7 is the most common serotype of Shiga-toxin-producing E. coli. Traditional methods for detecting E. coli O157:H7 are expensive, time-consuming, and less sensitive. A method with high sensitivity and high-resolution optical detection is utilizes the LSPR property of spherical gold nanoparticles (GNP). In this work, we constructed a novel nano-bio probe to detect E. coli O157:H7 by synthesizing citrate gold nanoparticle conjugated (non-covalent bond) with specific chicken anti-E. coli O157:H7 antibody (IgY) by changing the pH of the nanoparticles’ environment. UV-visible and DLS methods were used to confirm the bonding between the antibody and nanoparticles and the LSPR sensitivity of the nano-bio probe was evaluated by ELISA method. We could optically detect this bacterium in less than 2 h by measuring the LSPR band λ max shifts of GNPs. The sensitivity of this novel biosensor was determined by about 10 CFU/ml, using the LSPR property of spherical gold nanoparticles. So that, the LSPR λ max red shifted from 530 to 543 nm in presence of 10 CFU bacterium. In conclusion, this nano biosensor can be used to detect this important pathogen among the clinical specimens.

     

Neonatal oxytocin treatment modulates oxytocin receptor, atrial natriuretic peptide, nitric oxide synthase and estrogen receptor mRNAs expression in rat heart

Oxytocin (OT) has been implicated in reproductive functions, induction of maternal behavior as well as endocrine and neuroendocrine regulation of the cardiovascular system. Here we demonstrate that neonatal manipulation of OT can modulate the mRNAs expression for OT receptor (OTR), atrial natriuretic peptide (ANP), endothelial nitric oxide synthase (eNOS) and estrogen receptor alpha (ERalpha) in the heart. On the first day of postnatal life, female and male rats were randomly assigned to receive one of the following treatments: (a) 50microl i.p. injection of 7microg OT; (b) 0.7microg of OT antagonist (OTA); or (c) isotonic saline (SAL). Hearts were collected either on postnatal day 1 or day 21 (D1 or D21) and the mRNAs expression of OTR, ANP, inducible NOS (iNOS), eNOS, ERalpha and estrogen receptor beta (ERbeta) were compared by age, treatment, and sex utilizing real time PCR. OT treatment significantly increased heart OTR, ANP and eNOS mRNAs expression on D1 in both males and females, ERalpha increased only in females. While there were significant changes in the relative expression of all types of mRNA between D1 and D21, there were no significant treatment effects observed in D21 animals. OTA treatment significantly decreased basal ANP and eNOS mRNAs expression on D1 in both sexes. The results indicate that during the early postnatal period OT can have an immediate effect on the expression OTR, ANP, eNOS, and ERalpha mRNAs and that these effects are mitigated by D21. Also with the exception of ERalpha mRNA, the effects are the same in both sexes.     

Short-term changes in prostacyclin secretory profile of irradiated rat cervical spinal cord

Prostaglandins changes in radiation myelopathy (RM) have been previously reported. In the present study, we decided to determine the profile of Prostacyclin (PGI2) content in irradiated rat cervical cord. Wistar rats were irradiated with doses of 2,4,6,15,25 and 30 Gy of X-rays. After 24 h, 2 and 13 weeks post-irradiation, samples of spinal cord were prepared for evaluation of PGI2 and histopathologic changes. Prostacyclin content was determined by quantification of 6-keto-prostaglandin-F1alpha (prostacyclin major metabolite). Irradiated segments of spinal cord were stained routinely for histological studies. Results of irradiated were compared to control groups. Average ratio values of 6-keto-PG-F1alpha for doses of 2-30 Gy were between 67.5% and 107%, 65.41% and 100.54%, and 62.20% and 98.89% for 24 h, 2 and 13 weeks post-irradiation, respectively. Histopathological studies showed marked gliosis and vascularities in irradiated specimens. PGI2 bimodal secretory profile was observed along with histopathological changes in this study. Our results can further emphasize on the role of PGI2 in RM.     

Mixed Mating System Are Regulated by Fecundity in Shorea curtisii (Dipterocarpaceae) as Revealed by Comparison under Different Pollen Limited Conditions

The maintenance of mixed mating was studied in Shorea curtisii, a dominant and widely distributed dipterocarp species in Southeast Asia. Paternity and hierarchical Bayesian analyses were used to estimate the parameters of pollen dispersal kernel, male fecundity and self-pollen affinity. We hypothesized that partial self incompatibility and/or inbreeding depression reduce the number of selfed seeds if the mother trees receive sufficient pollen, whereas reproductive assurance increases the numbers of selfed seeds under low amounts of pollen. Comparison of estimated parameters of self-pollen affinity between high density undisturbed and low density selectively logged forests indicated that self-pollen was selectively excluded from mating in the former, probably due to partial self incompatibility or inbreeding depression until seed maturation. By estimating the self-pollen affinity of each mother tree in both forests, mother trees with higher amount of self-pollen indicated significance of self-pollen affinity with negative estimated value. The exclusion of self-fertilization and/or inbreeding depression during seed maturation occurred in the mother trees with large female fecundity, whereas reproductive assurance increased self-fertilization in the mother trees with lower female fecundity.