Complementary Fourier Single-Pixel Imaging.

Single-pixel imaging, with the advantages of a wide spectrum, beyond-visual-field imaging, and robustness to light scattering, has attracted increasing attention in recent years. Fourier single-pixel imaging (FSI) can reconstruct sharp images under sub-Nyquist sampling. However, the conventional FSI has difficulty balancing imaging quality and efficiency. To overcome this issue, we proposed a novel approach called complementary Fourier single-pixel imaging (CFSI) to reduce the number of measurements while retaining its robustness. The complementary nature of Fourier patterns based on a four-step phase-shift algorithm is combined with the complementary nature of a digital micromirror device. CFSI only requires two phase-shifted patterns to obtain one Fourier spectral value. Four light intensity values are obtained by loading the two patterns, and the spectral value is calculated through differential measurement, which has good robustness to noise. The proposed method is verified by simulations and experiments compared with FSI based on two-, three-, and four-step phase shift algorithms. CFSI performed better than the other methods under the condition that the best imaging quality of CFSI is not reached. The reported technique provides an alternative approach to realize real-time and high-quality imaging.

Systematic Design of a Quorum Sensing-Based Biosensor for Enhanced Detection of Metal Ion in Escherichia Coli.

With the recent industrial expansion, heavy metals and other pollutants have increasingly contaminated our living surroundings. The non-degradability of heavy metals may lead to accumulation in food chains and the resulting toxicity could cause damage in organisms. Hence, detection techniques have gradually received attention. In this study, a quorum sensing (QS)-based amplifier is introduced to improve the detection performance of metal ion biosensing. The design utilizes diffusible signal molecules, which freely pass through the cell membrane into the environment to communicate with others. Bacteria cooperate via the cell-cell communication process, thereby displaying synchronous behavior, even if only a minority of the cells detect the metal ion. In order to facilitate the design, the ability of the engineered biosensor to detect metal ion is described in a steady state model. The design can be constructed according to user-oriented specifications by selecting adequate components from corresponding libraries, with the help of a genetic algorithm (GA)-based design method. The experimental results validate enhanced efficiency and detection performance of the quorum sensing-based biosensor of metal ions.

Transforming growth factor-beta regulation of the insulin-like growth factor binding protein-4 protease system in cultured human osteoblasts.

IGFBP-4 is an inhibitor of IGF-I in bone. We show that TGF-beta regulates IGFBP-4 and enhances IGF-I-stimulated growth of cultured human bone cells through increased expression of an IGFBP-4 protease, PAPP-A. This effect of TGF-beta on IGF-I bioavailability may promote local bone formation. Insulin-like growth factor binding protein (IGFBP-4) proteolysis is implicated in the regulation of local insulin-like growth factor (IGF)-I bioavailability during bone remodeling. The IGFBP-4 protease secreted by normal adult human osteoblastic (hOB) cells in culture is a novel metalloproteinase, pregnancy-associated plasma protein-A (PAPP-A). We have recently identified an inhibitor of PAPP-A, the precursor form of major basic protein (proMBP). Very little is known about the molecular regulation of this IGFBP-4 protease system. In the present study, we determined the effect of transforming growth factor (TGF)-beta and IGF-II, the two most abundant growth factors in human bone, on PAPP-A and proMBP expression in primary cultures of hOB cells. Treatment with TGF-beta resulted in time- and dose-dependent increases in PAPP-A mRNA expression, with a maximal 12-fold increase after 24 h of stimulation with 10 ng/ml TGF-beta. Increased PAPP-A levels in hOB cell-conditioned medium paralleled PAPP-A gene expression. In addition, TGF-beta completely suppressed proMBP expression. Treatment of hOB cells with IGF-II had no effect on PAPP-A or proMBP gene expression. However, IGFBP-4 proteolysis in cell-free assay was dependent on IGF-II, and there was increased IGF-II-dependent IGFBP-4 protease activity in conditioned medium from hOB cells that were treated with TGF-beta. IGF-I stimulation of hOB cell proliferation was markedly enhanced by pretreatment with TGF-beta and [Leu27]IGF-II, and this enhancement was prevented with protease-resistant IGFBP-4. In summary, TGF-beta regulates IGFBP-4 proteolysis in hOB cells through increased expression of the protease, PAPP-A, and decreased expression of the inhibitor, proMBP. However, functional activation of the IGFBP-4 protease system is dependent on IGF-II, which acts at a post-translational level. These data support a model whereby local TGF-beta and IGF-II in the bone microenvironment coordinately amplify IGF-I bioavailability through controlled IGFBP-4 proteolysis, which may be a means to promote bone formation.

Molecular regulation of the IGF-binding protein-4 protease system in human fibroblasts: identification of a novel inducible inhibitor.

The IGF-binding protein-4 (IGFBP-4) protease system is an important regulator of local IGF bioavailability and cell growth. Recently, the IGF-dependent IGFBP-4 protease secreted by cultured human fibroblasts was identified as pregnancy-associated plasma protein A (PAPP-A). In pregnancy serum, PAPP-A circulates as a disulfide-bound complex with the precursor form of major basic protein (pro-MBP), and in this complex PAPP-A's proteolytic activity is not evident. In this study we analyzed the IGFBP-4 protease system in normal human fibroblasts to determine regulation outside of pregnancy. Treatment with the phorbol ester tumor promoter, beta-phorbol 12,13-didecanoate (beta-PDD), resulted in time-dependent inhibition of the IGF-dependent IGFBP-4 protease activity in cell-conditioned medium, which was evident at 6 h and complete by 24 h. PAPP-A mRNA was constitutively expressed in control cells, and levels were decreased only after 24 h of beta-PDD treatment. Secretion of PAPP-A protein into conditioned medium did not change with beta-PDD treatment. On the other hand, pro-MBP mRNA was undetectable in control human fibroblasts, and treatment with beta-PDD induced pro-MBP mRNA and protein expression within 6 h. beta-PDD-induced pro-MBP mRNA expression and protease inhibition were blocked with an inhibitor of RNA synthesis, actinomycin D. Actinomycin D had no effect on PAPP-A mRNA levels in the absence or presence of beta-PDD. Similarly, transformation of human fibroblasts with simian virus 40 large T antigen resulted in the synthesis of pro-MBP mRNA and protein and inhibition of IGFBP-4 protease activity. Coculture of fibroblasts with cells transfected with pro-MBP cDNA resulted in inhibition of IGFBP-4 proteolytic activity without having any effect on PAPP-A synthesis. In summary, phorbol ester tumor promoters and simian virus 40 transformation regulate IGFBP-4 proteolysis in human fibroblasts through induction of a novel inhibitor of PAPP-A, pro-MBP. These findings expand our understanding of the IGFBP-4 protease system and suggest an additional level of local cell growth control.

Pregnancy-associated plasma protein a gene expression as a target of inflammatory cytokines.

Pregnancy-associated plasma protein A (PAPP-A) cleaves IGF-binding protein-4 (IGFBP-4) and appears to enhance local IGF bioavailability in response to injury. In this study we determined the effects of growth factors and cytokines involved in the healing process on PAPP-A expression in human dermal fibroblasts. There was no effect of platelet-derived growth factor, epidermal growth factor, or basic fibroblast growth factor on PAPP-A mRNA expression in these cells. However, treatment with the proinflammatory cytokines, TNFalpha and IL-1 beta, resulted in time- and dose-dependent increases in PAPP-A mRNA and protein expression (3- to 4-fold maximal effects), which were prevented by actinomycin D. On the other hand, interferon-gamma (IFN gamma) treatment markedly inhibited PAPP-A expression. IGFBP-4 proteolytic activity was increased 4-fold in medium from TNFalpha- and IL-1 beta-treated (1 nm) cells and decreased 40% in medium from IFN gamma-treated (1 nm) cells. IGF-I-stimulated [(3)H]thymidine incorporation was significantly enhanced by pretreatment with 1 nm TNFalpha, and this enhancement was blocked in the presence of protease-resistant IGFBP-4. In conclusion, PAPP-A expression is regulated by inflammatory cytokines in adult human fibroblasts, with functional consequences on IGFBP-4 protease activity and IGF-I bioavailability. These data provide a mechanism for the regulation of PAPP-A in response to injury and further implicate PAPP-A in the wound-healing processes.

Localization and regulation of pregnancy-associated plasma protein a expression in healing human skin.

Pregnancy-associated plasma protein A (PAPP-A) is an IGF-binding protein-4 (IGFBP-4) metalloproteinase that cleaves inhibitory IGFBP-4 to amplify local IGF-I bioavailability in vitro. Thus it has functional implications in injury/repair responses. In this study we determined PAPP-A expression in healing human skin. Wounds were induced with a scalpel on the forearms of three normal subjects and were allowed to heal by first intention. Biopsies obtained on d 0, 2, 8, and 14 were processed for immunohistochemical detection of PAPP-A, IGF-I, and IGFBP-4. In uninjured skin (d 0), strong staining for PAPP-A was present in the epidermis, sweat and sebaceous gland epithelial cells, hair follicles, and blood vessels; no PAPP-A was detected in dermal fibroblasts or with mature collagen bundles. IGF-I localized strongly to epithelial cells of skin glands was weak to moderate in epidermis and blood vessels, and was absent in dermal cells. Weak focal staining for IGFBP-4 was found within uninjured epidermis. During wound healing, PAPP-A expression was induced in dermal granulation tissue within and adjacent to the injury. PAPP-A was present in dermis on d 2 and was increased in intensity and extent on d 8 and 14. PAPP-A expression also increased in the epidermis. PAPP-A expression in cells of granulation tissue colocalized with alpha-smooth actin staining of myofibroblasts and new blood vessels as well as with CD68 staining of macrophages and was associated with the compact, newly synthesized collagen of the healing wound. IGF-I staining was enhanced in the epidermis localized to the area of the incision and in granulation tissue associated with lymphoid cells. IGFBP-4 staining of the epidermis remained unchanged during wound healing, but was induced in the fibroblastic cells of granulation tissue over time. These data demonstrate localized and regulated expression of PAPP-A in human skin and suggest that PAPP-A may play an important role in an integrated IGF system in wound healing and tissue remodeling in vivo.

Regulation of pregnancy-associated plasma protein-A expression in cultured human osteoblasts.

Pregnancy-associated plasma protein-A (PAPP-A) is a metalloproteinase secreted by cultured human osteoblasts that has been implicated in the regulation of local insulin-like growth factor (IGF) bioavailability during bone growth and remodeling. However, very little is known about the regulation of PAPP-A expression in bone. In this study, we determined the effect of systemic and local osteoregulatory factors on PAPP-A mRNA and protein expression in normal human osteoblasts (hOB cells). Treatment of hOB cells with particular peptide growth factors (basic fibroblast growth factor, epidermal growth factor), steroid hormones (dexamethasone, 1,25-dihydroxyvitamin D(3)), and cytokines [interleukin-6 (IL-6), IL-13, oncostatin M] with known involvement in bone cell physiology had no significant effect on PAPP-A expression. Agents that increase intracellular cyclic AMP (forskolin, prostaglandin E(2)) increased PAPP-A mRNA and protein expression approximately 3-fold. Tumor necrosis factor alpha (TNFalpha), IL-1beta, and IL-4 also increased PAPP-A expression 3- to 4-fold. Transforming growth factor beta (TGFbeta) was previously shown to stimulate PAPP-A expression in hOB cells. The effects of TGFbeta, TNFalpha, and IL-1beta were additive, whereas the effects of TGFbeta and IL-4 were synergistic. In summary, TNFalpha, IL-1beta, and IL-4 were identified as potent stimulators of PAPP-A expression in primary cultures of human osteoblasts. These findings suggest a mechanism whereby cytokines present in bone and bone marrow could augment IGF bioavailability during skeletal growth and remodeling.

Axon regeneration through scaffold into distal spinal cord after transection.

We employed Fast Blue (FB) axonal tracing to determine the origin of regenerating axons after thoracic spinal cord transection injury in rats. Schwann cell (SC)-loaded, biodegradable, poly(lactic-co-glycolic acid) (PLGA) scaffolds were implanted after transection. Scaffolds loaded with solubilized basement membrane preparation (without SCs) were used for negative controls, and nontransected cords were positive controls. One or 2 months after injury and scaffold implantation, FB was injected 0-15 mm caudal or about 5 mm rostral to the scaffold. One week later, tissue was harvested and the scaffold and cord sectioned longitudinally (30 microm) on a cryostat. Trans-scaffold labeling of neuron cell bodies was identified with confocal microscopy in all cell-transplanted groups. Large (30-50 microm diameter) neuron cell bodies were predominantly labeled in the ventral horn region. Most labeled neurons were seen 1-10 mm rostral to the scaffold, although some neurons were also labeled in the cervical cord. Axonal growth occurred bidirectionally after cord transection, and axons regenerated up to 14 mm beyond the PLGA scaffolds and into distal cord. The extent of FB labeling was negatively correlated with distance from the injection site to the scaffold. Electron microscopy showed myelinated axons in the transverse sections of the implanted scaffold 2 months after implantation. The pattern of myelination, with extracellular collagen and basal lamina, was characteristic of SC myelination. Our results show that FB labeling is an effective way to measure the origin of regenerating axons.

Pregnancy-associated plasma protein-A (PAPP-A) expression and insulin-like growth factor binding protein-4 protease activity in normal and malignant ovarian surface epithelial cells.

Pregnancy-Associated Plasma Protein-A (PAPP-A) proteolyses insulin-like growth factor binding protein-4 (IGFBP-4), thereby regulating local IGF availability. Reduced PAPP-A mRNA expression has been reported in ovarian cancer specimens compared to normal ovarian surface epithelial cells (OSE). To characterize PAPP-A expression and proteolytic activity in OSE, we developed a lifespan-extended human cell model using a temperature-sensitive mutant of the SV40 large T antigen (SV40LT). These OSE(tsT) cells proliferate at 34 degrees C (i.e., when SV40LT-positive), but not at 39 degrees C, a temperature at which the SV40LT is unstable (SV40LT-negative). Proteolysis of radiolabeled IGFBP-4 in conditioned media from OSE(tsT) lines was IGF-dependent and blocked by anti-PAPP-A antisera. Temperature shifts that eliminated stable SV40LT induced a 7-fold increase in PAPP-A mRNA and a 4-fold increase in protein. The converse experiment (shifting to SV40LT-positive conditions) resulted in decreased levels of PAPP-A mRNA but little change in PAPP-A protein. Nevertheless, there was a marked reduction in IGF-BP-4 proteolytic activity in medium of SV40LT-positive OSE-(tsT) cells. This decreased PAPP-A activity coincided with a nearly 20-fold increase in mRNA encoding a physiological inhibitor of PAPP-A, the precursor form of eosinophil Major Basic Protein (proMBP), and 4- to 5-fold increases in proMBP protein. Primary cultures of unmodified OSE expressed high levels of PAPP-A and undetectable proMBP, and therefore produced abundant IGFBP-4 protease activity. Short-term ovarian tumor cell cultures expressed variable levels of PAPP-A and high levels of proMBP, and consequently secreted little or no IGFBP-4 protease activity. The concurrent regulation of PAPP-A and its inhibitor, proMBP, suggests that IGFBP-4 proteolysis and local regulation of IGF availability may be altered in malignant ovarian epithelial cells.