hnRNP C and polypyrimidine tract-binding protein specifically interact with the pyrimidine-rich region within the 3'NTR of the HCV RNA genome

Like other members of the Flaviviridae family, the 3' non-translated region (NTR) of the hepatitis C virus (HCV) is believed to function in the initiation and regulation of viral RNA replication by interacting with components of the viral replicase complex. To inves-tigate the possibility that host components may also participate in this process, we used UV cross-linking assays to determine if any cellular proteins could bind specifically to the 3'NTR RNA. We demonstrate the specific interaction of two host proteins with the extensive pyrimidine-rich region within the HCV 3'NTR. One host protein migrates as a doublet with a molecular weight of 57 kDa and is immunoreactive with antisera specific for polypyrimidine tract-binding protein (PTB), and the other protein (35 kDa) is recognized by a monoclonal antibody specific for heterogeneous nuclear ribonucleoprotein C (hnRNP C). These results suggest that recognition of the large pyrimidine-rich region by PTB and hnRNP C may play a role in the initiation and/or regulation of HCV RNA replication.     

Molecular mechanisms involved in progesterone receptor regulation of uterine function

The ovarian steroid hormone progesterone is a major regulator of uterine function. The actions of this hormone is mediated through its cognate receptor, the progesterone receptor, Pgr. Ablation of the Pgr has shown that this receptor is critical for all female reproductive functions including the ability of the uterus to support and maintain the development of the implanting mouse embryo. High density DNA microarray analysis has identified direct and indirect targets of Pgr action. One of the targets of Pgr action is a member of the Hedgehog morphogen Indian Hedgehog, Ihh. Ihh and members of the Hh signaling cascade show a coordinate expression pattern in the mouse uterus during the preimplantation period of pregnancy. The expression of Ihh and its receptor Patched-1, Ptc1, as well as, down stream targets of Ihh-Ptch1 signaling, such as the orphan nuclear receptor COUP-TF II show that this morphogen pathway mediates communication between the uterine epithelial and stromal compartments. The members of the Ihh signaling axis may function to coordinate the proliferation, vascularization and differentiation of the uterine stroma during pregnancy. This analysis demonstrates that progesterone regulates uterine function in the mouse by coordinating the signals from the uterine epithelium to stroma in the preimplantation mouse uterus.     

Clinical Implications and Future Perspectives of Circulating Tumor Cells and Biomarkers in Clinical Outcomes of Colorectal Cancer

Colorectal cancer (CRC) is a major public health problem. Early CRC detection, pretherapeutic responsiveness prediction, and postoperative micrometastasis monitoring are the hallmarks for successful CRC treatment. Here, the methodologies used for detecting circulating tumor cells (CTCs) from CRC are reviewed. In addition to the traditional CRC biomarkers, the persistent presence of posttherapeutic CTCs indicates resistance to adjuvant chemotherapy and/or radiotherapy; hence, CTCs also play a decisive role in the subsequent relapse of CRC. Moreover, the genetic and phenotypic profiling of CTCs often differs from that of the primary tumor; this difference can be used to select the most effective targeted therapy. Consequently, studying CTCs can potentially individualize treatment strategies for patients with CRC. Therefore, CTC detection and characterization may be valuable tools for refining prognosis, and CTCs can be used in a real-time tumor biopsy for designing individually tailored therapy against CRC.     

Transforming growth factor-β1 suppresses hepatitis B virus replication by the reduction of hepatocyte nuclear factor-4α expression

Several studies have demonstrated that cytokine-mediated noncytopathic suppression of hepatitis B virus (HBV) replication may provide an alternative therapeutic strategy for the treatment of chronic hepatitis B infection. In our previous study, we showed that transforming growth factor-beta1 (TGF-β1) could effectively suppress HBV replication at physiological concentrations. Here, we provide more evidence that TGF-β1 specifically diminishes HBV core promoter activity, which subsequently results in a reduction in the level of viral pregenomic RNA (pgRNA), core protein (HBc), nucleocapsid, and consequently suppresses HBV replication. The hepatocyte nuclear factor 4alpha (HNF-4α) binding element(s) within the HBV core promoter region was characterized to be responsive for the inhibitory effect of TGF-β1 on HBV regulation. Furthermore, we found that TGF-β1 treatment significantly repressed HNF-4α expression at both mRNA and protein levels. We demonstrated that RNAi-mediated depletion of HNF-4α was sufficient to reduce HBc synthesis as TGF-β1 did. Prevention of HNF-4α degradation by treating with proteasome inhibitor MG132 also prevented the inhibitory effect of TGF-β1. Finally, we confirmed that HBV replication could be rescued by ectopic expression of HNF-4α in TGF-β1-treated cells. Our data clarify the mechanism by which TGF-β1 suppresses HBV replication, primarily through modulating the expression of HNF-4α gene.     

Resistin induces monocyte-endothelial cell adhesion by increasing ICAM-1 and VCAM-1 expression in endothelial cells via p38MAPK-dependent pathway

Resistin, firstly reported as an adipocyte-specific hormone, is suggested to be an important link between obesity and diabetes. Recent studies have suggested an association between resistin and atherogenic processes. The adhesion of circulating monocytes to endothelial cells is a critical step in the early stages of atherosclerosis. The purpose of the present study was to investigate the effect of resistin on the adhesion of THP-1 monocytes to human umbilical vein endothelial cells (HUVECs) and the underlying mechanism. Our results showed that resistin caused a significant increase in monocyte adhesion. In exploring the underlying mechanisms of resistin action, we found that resistin-induced monocyte adhesion was blocked by inhibition of p38MAPK activation using SB203580 and SB202190. Furthermore, resistin increased the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) by HUVECs and these effects were also p38MAPK-dependent. Resistin-induced monocyte adhesion was also blocked by monoclonal antibodies against ICAM-1 and VCAM-1. Taken together, these results show that resistin increases both the expression of ICAM-1 and VCAM-1 by endothelial cells and monocyte adhesion to HUVECs via p38MAPK-dependent pathways.     

Kinetics of tributyrin hydrolysis by lipase

The kinetics for the tributyrin hydrolysis using lipase (Pseudomonas fluorscenes CCRC-17015) were investigated in the liquid–liquid and liquid–solid–liquid reaction systems in a batch reactor. The lipase was covalently immobilized onto the surface of porous polymethylacrylamide (PMAA) crosslinking with N,N-methylene biacrylamide with a spacer of ethylenediamine actived by glutaraldehyde. The conditions such as tributyrin concentration, temperature, agitation, and pH value, were evaluated to achieve the optimum reaction conditions for both free lipase and immobilized lipase. The kinetic parameters in the reaction system were also obtained for two reaction systems. The turnover numbers calculated for free lipase and immobilized lipase were 29 and 5.7 s⁻¹, respectively. The parameters of k and k_m obtained using Lineweaver-Burk plot method were 26.2 mol/(mg min) and 1.35 mol/dm³ for free lipase, 5.2 mol/(mg min) and 0.2 mol/dm³ for immobilized lipase, respectively. The experimental results revealed good thermal stability, with greater stability at higher pH value for immobilized lipase in the liquid–solid–liquid reaction.     

Conjugation of Multiple Copies of Polyethylene Glycol to Hemoglobin Facilitated Through Thiolation: Influence on Hemoglobin Structure and Function

PEGylation induced changes in molecular volume and solution properties of HbA have been implicated as potential modulators of its vasoconstrictive activity. However, our recent studies with PEGylated Hbs carrying two PEG chains/Hb, have demonstrated that the modulation of the vasoconstrictive activity of Hb is not a direct correlate of the molecular volume and solution properties of the PEGylated Hb and implicated a role for the surface charge and/or the pattern of surface decoration of Hb with PEG. HbA has now been modified by thiolation mediated maleimide chemistry based PEGylation that does not alter its surface charge and conjugates multiple copies of PEG5K chains. This protocol has been optimized to generate a PEGylated Hb, (SP-PEG5K)₆-Hb, that carries ~six PEG5K chains/Hb – HexaPEGylated Hb. PEGylation increased the O₂ affinity of Hb and desensitized the molecule for the influence of ionic strength, pH, and allosteric effectors, presumably a consequence of the hydrated PEG-shell generated around the protein. The total PEG mass in (SP-PEG5K)₆-Hb, its molecular volume, O₂ affinity and solution properties are similar to that of another PEGylated Hb, (SP-PEG20K)₂-Hb, that carries two PEG20K chains/Hb. However, (SP-PEG5K)₆-Hb exhibited significantly reduced vasoconstriction mediated response than (SP-PEG20K)₂-Hb. These results demonstrate that the enhanced molecular size and solution properties achieved through the conjugation of multiple copies of small PEG chains to Hb is more effective in decreasing its vasoconstrictive activity than that achieved through the conjugation of a comparable PEG mass using a small number of large PEG chains.     

Life cycles,phenology and genetic structure of endangered Megacrania tsudai Shiraki (Phasmatodea: Phasmatidae): Male individuals from a geographic parthenogenesis species

Megacrania tsudai, a peripherally distributed member of Megacrania, requires conservation in Taiwan; it has limited distribution in Taiwan and its eastern offshore islands. It feeds on screw pines (Pandanus odoratissimus) in nature and has demonstrated a specific defensive mechanism involving actinidine secretion from the prothoracic gland. However, details of its distribution area, life cycle and developmental phenology remain largely unknown. In this study, a field survey and review of published works revealed M. tsudai distribution in coastal zones and along river shores near estuaries. At room temperature, the egg period was 128 days. The development of the first to sixth instars required 17, 26, 27, 26, 34 and 43 days, respectively, on average; and a generation cycle required approximately 204 days. The phenology of the mesonotal granules was recorded. Moreover, genetic analysis of the mitochondrial cytochrome oxidase I (COI), 16S ribosomal DNA (16S rDNA) and the nuclear ribosomal spacer indicated the occurrence of genetic drift. Therefore, the rearing procedures proposed in this study for the primary and last instars of M. tsudai can facilitate its conservation. Megacrania tsudai was previously recorded as parthenogenetic; however, two male individuals were fostered unexpectedly. The male body length was 91 mm, which is shorter than the female length (120 mm). During mating, the male climbs onto the female's back and protrudes its genitalia downward. Geographical parthenogenesis is likely the reproductive strategy among peripheral M. tsudai; however, the rarely found M. tsudai male could be an intermediate link of reproductive strategy in the transition from tychoparthenogenesis to parthenogenesis.