Nitric Oxide Inhibits Vascular Smooth Muscle Cell Proliferation and Neointimal Hyperplasia by Increasing the Ubiquitination and Degradation of UbcH10

Nitric oxide (NO) limits formation of neointimal hyperplasia in animal models of arterial injury in large part by inhibiting vascular smooth muscle cell (VSMC) proliferation through cell cycle arrest. The ubiquitin-conjugating enzyme UbcH10 is responsible for ubiquitinating cell cycle proteins for proper exit from mitosis. We hypothesize that NO prevents VSMC proliferation, and hence neointimal hyperplasia, by decreasing levels of UbcH10. Western blotting and immunofluorescent staining showed that NO reduced UbcH10 levels in a concentration-dependent manner in VSMC harvested from the abdominal aortas of Sprague-Dawley rats. Treatment with NO or siRNA to UbcH10 decreased both UbcH10 levels and VSMC proliferation (P<0.001), while increasing UbcH10 levels by plasmid transfection or angiotensin II stimulation increased VSMC proliferation to 150% (P=0.008) and 212% (P=0.002) of control, respectively. Immunofluorescent staining of balloon-injured rat carotid arteries showed a ~4-fold increase in UbcH10 levels, which was profoundly decreased following treatment with NO. Western blotting of carotid artery lysates showed no UbcH10 in uninjured vessels, a substantial increase in the injury alone group, and a significant decrease in the injury+NO group (~3-fold reduction versus injury alone). Importantly, in vitro and in vivo, a marked increase in polyubiquitinated UbcH10 was observed in the NO-treated VSMC and carotid arteries, respectively, indicating that NO may be decreasing unmodified UbcH10 levels by increasing its ubiquitination. Central to our hypothesis, we report that NO decreases UbcH10 levels in VSMC in vitro and following arterial injury in vivo in association with increasing polyubiquitinated-UbcH10 levels. These changes in UbcH10 levels correlate with VSMC proliferation and neointimal hyperplasia, making UbcH10 a promising therapeutic target for inhibiting this proliferative disease.     

Rapid determination of gatifloxacin in biological samples and pharmaceutical products using europium‐sensitized fluorescence spectrophotometry

A europium‐sensitized fluorescence spectrophotometry method using an anionic surfactant, sodium dodecyl benzene sulphonate (SDBS), was developed for the determination of gatifloxacin (GFLX). The GFLX–Eu³⁺–SDBS system was studied and it was found that SDBS significantly enhanced the fluorescence intensity of the GFLX–Eu³⁺ complex (about 25‐fold). The optimal experimental conditions were determined as follows: excitation and emission wavelengths of 338 and 617 nm, pH 7.5, 3.0 × 10^–6 mol/L europium(III), and 5.0 × 10^–5 mol/L SDBS. The enhanced fluorescence intensity of the system (ΔI_f) showed a good linear relationship with the concentration of GFLX over the range 1.0 × 10^–8–8.0 × 10^–7 mol/L with a correlation coefficient of 0.9990. The detection limit (S:N = 3) was determined as 1.0 × 10^–9 mol/L. This method has been successfully applied for the determination of GFLX in pharmaceuticals and human urine/serum samples. Compared with most other methods reported, the rapid and simple procedure proposed here offered higher sensitivity, wider linear range and good stability. The luminescence mechanism of the system is also discussed in detail. Copyright © 2008 John Wiley & Sons, Ltd.     

Is food resource partitioning responsible for deviation of echolocation call frequencies from allometry in <Emphasis Type="Italic">Rhinolophus macrotis</Emphasis>?

There is an apparent allometric relationship between peak frequency of echolocation and body size in rhinophilids. However, some rhinolophids deviate from this rule. To date this variation has been explained as a result of partitioning of communication channels. An alternative hypothesis that food resource partitioning results in this divergence in expected frequencies was tested by comparing prey selection between Rhinolophus macrotis Blyth, 1844 and Rhinolophus lepidus Blyth, 1844 in Yunnan Province, China. These two sympatric species are morphologically similar but acoustically divergent: R. macrotis has an echolocation frequency significantly lower than that predicted by the allometric relationship, whereas that of R. lepidus agreed with expectations. Prey selection experiments, conducted in a flight tent, indicated that the dominant prey taxa of R. macrotis were Lasiocampidae, Arctiidae and Noctuidae, whilst that of R. lepidus were Arctiidae, Noctuidae and Ichneumonidae. R. macrotis ate more earless moths and fewer eared moths than R. lepidus, and R. macrotis fed on larger prey in general and captured a wider size range than that captured by R. lepidus. These results confirmed the existence of finely tuned trophic niche differentiation and suggested that food resource partitioning is one of the factors leading to lower peak frequency of calls in R. macrotis.     

A20 plays a critical role in the immunoregulatory function of mesenchymal stem cells

Mesenchymal stem cells (MSCs) possess an immunoregulatory capacity and are a therapeutic target for many inflammation‐related diseases. However, the detailed mechanisms of MSC‐mediated immunosuppression remain unclear. In this study, we provide new information to partly explain the molecular mechanisms of immunoregulation by MSCs. Specifically, we found that A20 expression was induced in MSCs by inflammatory cytokines. Knockdown of A20 in MSCs resulted in increased proliferation and reduced adipogenesis, and partly reversed the suppressive effect of MSCs on T cell proliferation in vitro and inhibited tumour growth in vivo. Mechanistic studies indicated that knockdown of A20 in MSCs inhibited activation of the p38 mitogen‐activated protein kinase (MAPK) pathway, which potently promoted the production of tumour necrosis factor (TNF)‐α and inhibited the production of interleukin (IL)‐10. Collectively, these data reveal a crucial role of A20 in regulating the immunomodulatory activities of MSCs by controlling the expression of TNF‐α and IL‐10 in an inflammatory environment. These findings provide novel insights into the pathogenesis of various inflammatory‐associated diseases, and are a new reference for the future development of treatments for such afflictions.     

Amplification, analysis and chromosome mapping of novel homeobox-containing and homeobox-flanking sequences in rice

Homeobox genes, widely distributed among animal and plant kingdoms, play an important role in developmental process. Several homeobox conserved fragments were amplified by PCR and the flanking regions were also obtained by an LM-PCR procedure. Sequencing and Southern analysis showed that they belong to a homeobox gene family of rice. Six homeobox-containing fragments were mapped on the molecular linkage map of rice. They were located on chromosomes 3, 4 and 7 respectively. It is noteworthy that there are 4 homeobox fragments located on rice chromosome 3 and the result is also consistent with the comparative genomics between rice and maize.     

Alpha-melanocyte stimulating hormone protects retinal pigment epithelium cells from oxidative stress through activation of melanocortin 1 receptor–Akt–mTOR signaling

Patients with age related macular degeneration (AMD) will develop vision loss in the center of the visual field. Reactive oxygen species (ROS)-mediated retinal pigment epithelium (RPE) cell apoptosis is an important contributor of AMD. In this study, we explored the pro-survival effect of α-melanocyte stimulating hormone (α-MSH) on oxidative stressed RPE cells. We found that α-MSH receptor melanocortin 1 receptor (MC1R) was functionally expressed in primary and transformed RPE cells. RPE cells were response to α-MSH stimulation. α-MSH activated Akt/mammalian target of rapamycin (mTOR) and Erk1/2 signalings in RPE cells, which were inhibited by MC1R siRNA knockdown. α-MSH protected RPE cells from hydrogen peroxide (H₂O₂)-induced apoptosis, an effect that was almost abolished when MC1R was depleted by siRNA. α-MSH-mediated S6K1 activation and pro-survival effect against H₂O₂ was inhibited by Akt inhibitors (perifosine, MK-2206 and LY294002). Further, mTOR inhibition by rapamycin, or by mTOR siRNA knockdown, diminished α-MSH’s pro-survival effect in RPE cells. Thus, Akt and its downstream mTOR signaling mediates α-MSH-induced survival in RPE cells. In summary, we have identified a new α-MSH–MC1R physiologic pathway that reduces H₂O₂-induced RPE cell damage, and might minimize the risk of developing AMD.     

Apoptosis induced by (di-isopropyloxyphoryl-Trp)2-Lys-OCH3 in K562 and HeLa cells

According to the method used in our laboratory, our group synthesized (DIPP-Trp)₂-Lys-OCH₃. It inhibited the proliferation of K562 and HeLa cells in a dose-and time-dependent manner with an IC₅₀ of 15.12 and 42.23 µM, respectively. (DIPP-Trp)₂-Lys-OCH₃ induced a dose-dependent increase of the G2/M cell population in K562 cells, and S cell population in HeLa cells; the sub-G0 population increased dramatically in both cell lines as seen by PI staining experiments using a FACS Calibur Flow cytometer (BeckmanCoulter, USA). Phosphatidylserine could significantly translocate to the surface of the membrane in (DIPP-Trp)₂-Lys-OCH₃-treated K562 and HeLa cells. The increase of an early apoptotic population was observed in a dose-dependent manner by both annexin-FITC and PI staining. It was concluded that (DIPP-Trp)₂-Lys-OCH₃ not only induced cells to enter into apoptosis, but also affected the progress of the cell cycle. It may have arrested the K562 and HeLa cells in the G₂/M, S phases, respectively. The apoptotic pathway was pulsed at this point, resulting in the treated cells entering into programmed cell death. (DIPP-Trp)₂-Lys-OCH₃ is a potential anticancer drug that intervenes in the signalling pathway.     

Challenge of weed risk assessment (WRA) for ecological restoration in China: The case of Rhus typhina L. and the new officially released weed risk assessment system

China has conducted many ecological restoration projects to alleviate environmental degradation, with numerous alien species having good adaptability widely used for fast effect. However, unsuitable plant selection may bring negative impacts and even cause biological invasion. Weed risk assessment (WRA) is therefore indispensable, yet it is often contentious especially concerning those species that has been widely utilized before weedy attributes were noted. In this paper, we take Rhus typhina L. as an example to illustrate how scientific researches and social factors can influence the evaluation of alien species and cause challenge to WRA. The evaluation of this species is not only a pure scientific issue but also influenced by many factors such as the introduction history, current utilization status in afforestation, and divergent underlying values varying through persons and sectors. These factors determined the kind of data to be assessed, which lead to different evaluations, therefore, causing challenge to its WRA. We further examined newly officially released WRA system in China from the invasion biology viewpoint, arguing that it had some major flaws in design and validation and need much improvement. Considering the human dimension and biological characters together, we suggest that the “black list” and “green list” approaches with rigorous expert assessment should be adopted simultaneously in alien species management in China.     

RETRACTED ARTICLE: Therapeutic DNA Vaccination as a Repair Strategy Following Spinal Cord Injury

Myelin-derived proteins, such as tenascin-R (TN-R), myelin associate glycoprotein (MAG), oligodendrocyte-myelin glycoprotein (OMgp), and Nogo-A, inhibit the central nervous system regeneration. In this study, the DNA vaccine encoding for oligodendrocyte and myelin-related antigens was employed to attenuate the axonal growth inhibitory properties of myelin in the setting of spinal cord injury. Using a rat spinal cord dorsal hemisection model, the vaccine directed against the inhibitory epitopes of Nogo-A, MAG, OMgp, and TN-R was administered intramuscularly once a week following spinal cord injury, supplemented with local application of specific anti-sera against the four antigens. Anterograde labeling of dorsal column fibers showed active axonal regeneration through the lesion site at the eighth week following the treatment in experimental group but not in control groups. Light microscopic and ultrastructural analysis revealed that vaccination with these myelin-related antigens did not lead to demyelinating disease. OMgp and TN-R levels were down-regulated at the lesion site together with a parallel increase in growth-associated protein 43 levels in the treatment groups. This study reveals the effective approach of a DNA vaccine strategy by attaining the special antibody to direct neutralization of the myelin inhibitors during spinal cord injury.     

Cyanidin-3-glucoside, a natural product derived from blackberry, exhibits chemopreventive and chemotherapeutic activity

Epidemiological data suggest that consumption of fruits and vegetables has been associated with a lower incidence of cancer. Cyanidin-3-glucoside (C3G), a compound found in blackberry and other food products, was shown to possess chemopreventive and chemotherapeutic activity in the present study. In cultured JB6 cells, C3G was able to scavenge ultraviolet B-induced *OH and O2-* radicals. In vivo studies indicated that C3G treatment decreased the number of non-malignant and malignant skin tumors per mouse induced by 12-O-tetradecanolyphorbol-13-acetate (TPA) in 7,12-dimethylbenz[a]anthracene-initiated mouse skin. Pretreatment of JB6 cells with C3G inhibited UVB- and TPA-induced transactivation of NF-kappaB and AP-1 and expression of cyclooxygenase-2 and tumor necrosis factor-alpha. These inhibitory effects appear to be mediated through the inhibition of MAPK activity. C3G also blocked TPA-induced neoplastic transformation in JB6 cells. In addition, C3G inhibited proliferation of a human lung carcinoma cell line, A549. Animal studies showed that C3G reduced the size of A549 tumor xenograft growth and significantly inhibited metastasis in nude mice. Mechanistic studies indicated that C3G inhibited migration and invasion of A549 tumor cells. These finding demonstrate for the first time that a purified compound of anthocyanin inhibits tumor promoter-induced carcinogenesis and tumor metastasis in vivo.