重组骆驼蓬脂转移蛋白对宫颈癌HeLa细胞化疗的增敏作用

[目的]研究重组骆驼蓬脂转移蛋白(recombinant Peganum harmala lipid transfer protein,r Ph LTP)与顺铂(DDP)协同对宫颈癌He La细胞的抑制作用及可能的作用机制。[方法]四甲基偶氮咗蓝(MTT)法检测He La细胞生长的抑制率;Annexin-V/PI双染法检测细胞凋亡率,流式细胞术检测细胞内活性氧(ROS)的变化情况。[结果]r Ph LTP与DDP联合应用后与相应的DDP组相比,对细胞生长的抑制率显著增强(P0.01),低浓度r Ph LTP联合2.5μg/m L顺铂对He La细胞的抑制率为(42.50±0.059)%,远高于高浓度顺铂(10μg/m L)对He La细胞的抑制率(36.88±0.134)%,细胞的凋亡率显著增加(P0.01),细胞内ROS极明显升高(P0.001)。[结论]r Ph LTP能够显著增强顺铂对He La细胞的化疗敏感性(P0.001),这种作用可能与r Ph LTP增强顺铂诱导细胞内活性氧的产生导致细胞凋亡有关。     

Variations in sexual and asexual reproduction of Scirpus mariqueter along an elevational gradient

In order to assess the importance of sexual and asexual reproduction during the life history of Scirpus mariqueter, its reproductive and growth characters were concurrently examined along an elevational gradient (from low elevation to high elevation). The proportions of flowering shoot and inflorescence mass, seed : flower ratio and seed weight were used to quantify the investment in sexual reproduction. The proportions of current-year shoot and rhizome mass were used to quantify the investment in asexual reproduction, and the proportion of corm mass was used for growth, respectively. It was found that vegetative propagation predominated at low elevation, whereas sexual reproduction predominated at high elevation; and that sexual reproduction increased with declining asexual reproduction along the gradient. The results suggest that asexual reproduction is relatively favored in the early life stage, whereas sexual reproduction is favored when the population becomes mature and aged, probably because of the functional differentiation between the two reproductive types. Sexual productive characters (i.e. the proportions of flowering shoot and inflorescence mass) were negatively correlated to both growth and asexual reproductive characters along the gradient, indicating there might exist some trade-offs among growth, sexual and asexual reproduction during the life history. However, no obvious pattern was found between asexual reproductive characters and growth characters along the elevational gradient, possibly because of the varied relationships between them at different life stages. The variations in sexual and asexual reproduction in the species and the relationship between them are thought to be of great significance for local population growth, species persistence and evolution.     

Immunohistochemical analysis of pRb2/p130, VEGF, EZH2, p53, p16(INK4A), p27(KIP1), p21(WAF1), Ki-67 expression patterns in gastric cancer

Although the considerable progress against gastric cancer, it remains a complex lethal disease defined by peculiar histological and molecular features. The purpose of the present study was to investigate pRb2/p130, VEGF, EZH2, p53, p16(INK4A), p27(KIP1), p21(WAF1), Ki-67 expressions, and analyze their possible correlations with clinicopathological factors. The expression patterns were examined by immunohistochemistry in 47 patients, 27 evaluated of intestinal-type, and 20 of diffuse-type, with a mean follow up of 56 months and by Western blot in AGS, N87, KATO-III, and YCC-2, -3, -16 gastric cell lines. Overall, stomach cancer showed EZH2 correlated with high levels of p53, Ki-67, and cytoplasmic pRb2/p130 (P < 0.05, and P < 0.01, respectively). Increased expression of EZH2 was found in the intestinal-type and correlated with the risk of distant metastasis (P < 0.05 and P < 0.01, respectively), demonstrating that this protein may have a prognostic value in this type of cancer. Interestingly, a strong inverse correlation was observed between p27(KIP1) expression levels and the risk of advanced disease and metastasis (P < 0.05), and a positive correlation between the expression levels of p21(WAF1) and low-grade (G1) gastric tumors (P < 0.05), confirming the traditionally accepted role for these tumor-suppressor genes in gastric cancer. Finally, a direct correlation was found between the expression levels of nuclear pRb2/p130 and low-grade (G1) gastric tumors that was statistically significant (P < 0.05). Altogether, these data may help shed some additional light on the pathogenetic mechanisms related to the two main gastric cancer histotypes and their invasive potentials.     

Two novel ubiquitin-fold modifier 1 (Ufm1)-specific proteases, UfSP1 and UfSP2

Ubiquitin-fold modifier 1 (Ufm1) is a recently identified new ubiquitin-like protein, whose tertiary structure displays a striking resemblance to ubiquitin. Similar to ubiquitin, it has a Gly residue conserved across species at the C-terminal region with extensions of various amino acid sequences that need to be processed in vivo prior to conjugation to target proteins. Here we report the isolation, cloning, and characterization of two novel mouse Ufm1-specific proteases, named UfSP1 and UfSP2. UfSP1 and UfSP2 are composed of 217 and 461 amino acids, respectively, and they have no sequence homology with previously known proteases. UfSP2 is present in most, if not all, of multicellular organisms including plant, nematode, fly, and mammal, whereas UfSP1 could not be found in plant and nematode upon data base search. UfSP1 and UfSP2 cleaved the C-terminal extension of Ufm1 but not that of ubiquitin or other ubiquitin-like proteins, such as SUMO-1 and ISG15. Both were also capable of releasing Ufm1 from Ufm1-conjugated cellular proteins. They were sensitive to inhibition by sulfhydryl-blocking agents, such as N-ethylmaleimide, and their active site Cys could be labeled with Ufm1-vinylmethylester. Moreover, replacement of the conserved Cys residue by Ser resulted in a complete loss of the UfSP1 and UfSP2 activities. These results indicate that UfSP1 and UfSP2 are novel thiol proteases that specifically process the C terminus of Ufm1.     

Landscape mulches and termite nutritional ecology: growth and survival of incipient colonies of Coptotermes formosanus (Isoptera: Rhinotermitidae)

Alate swarms are one of the major visible signs of the expansion of the Formosan subterranean termite, Coptotermes formosanus Shiraki (Isoptera: Rhinotermitidae), in an area. Successful establishment of an incipient colony is thought to mainly rely on available food resources and moisture. The large-scale use of tree-based mulches in landscapes may inadvertently contribute to local establishment and growth of C. formosanus colonies. This research investigated the nutritional ecology of incipient colonies of C. formosanus feeding on seven tree-based, weathered, and nonweathered landscape mulches: pine straw, pine bark, cedar wood, water oak, eucalyptus, cypress, and melaleuca. Incipient colonies of C. formosanus feeding on pine straw, either weathered or nonweathered, produced significantly more progeny over the course of 1-yr feeding than colonies feeding on the other mulches tested. Regardless of weathered or not, the incipient colonies feeding on pine straw, eucalyptus, bald cypress, and water oak mulches had significantly greater survival rates after 360 d (53-77%) than colonies feeding on the other mulches tested (0-13%), but colonies feeding on nonweathered water oak had significantly lower survival (8%) than those kept on weathered water oak (58%). Colony fitness values were significantly different between the weathering treatment groups and among the different types of mulches. With regard to colony growth characteristics, three distinct growth patterns were identified: a high number of progeny (>100) with high colony survival rate (>50%), a medium number of progeny (12-50) with high colony survival rate (>50%), and a small number of progeny (0-10) with low colony survival rate (<5%). These findings suggest that different types of mulch substrates could significantly impact the nutritional ecology of the founding pairs and the successful establishment of incipient colonies during the swarming season.     

Sucrose nonfermenting AMPK-related kinase (SNARK) regulates exercise-stimulated and ischemia-stimulated glucose transport in the heart

The signaling mechanisms mediating myocardial glucose transport are not fully understood. Sucrose nonfermenting AMP-activated protein kinase (AMPK)-related kinase (SNARK) is an AMPK-related protein kinase that is expressed in the heart and has been implicated in contraction-stimulated glucose transport in mouse skeletal muscle. We first determined if SNARK is phosphorylated on Thr²⁰⁸, a site critical for SNARK activity. Mice were treated with exercise, ischemia, submaximal insulin, or maximal insulin. Treadmill exercise slightly, but significantly increased SNARK Thr²⁰⁸ phosphorylation. Ischemia also increased SNARK Thr²⁰⁸ phosphorylation, but there was no effect of submaximal or maximal insulin. HL1 cardiomyocytes were used to overexpress wild-type (WT) SNARK and to knockdown endogenous SNARK. Overexpression of WT SNARK had no effect on ischemia-stimulated glucose transport; however, SNARK knockdown significantly decreased ischemia-stimulated glucose transport. SNARK overexpression or knockdown did not alter insulin-stimulated glucose transport or glycogen concentrations. To study SNARK function in vivo, SNARK heterozygous knockout mice (SNARK^+/−) and WT littermates performed treadmill exercise. Exercise-stimulated glucose transport was decreased by ~50% in hearts from SNARK^+/− mice. In summary, exercise and ischemia increase SNARK Thr²⁰⁸ phosphorylation in the heart and SNARK regulates exercise-stimulated and ischemia-stimulated glucose transport. SNARK is a novel mediator of insulin-independent glucose transport in the heart.     

MicroRNA-323-3p promotes myogenesis by targeting Smad2

Skeletal muscle is an important and complex organ with multiple biological functions in humans and animals. Proliferation and differentiation of myoblasts are the key steps during the development of skeletal muscle. MicroRNA (miRNA) is a class of 21-nucleotide noncoding RNAs regulating gene expression by combining with the 3′-untranslated region of target messenger RNA. Many studies in recent years have suggested that miRNAs play a critical role in myogenesis. Through high-throughput sequencing, we found that miR-323-3p showed significant changes in the longissimus dorsi muscle of Rongchang pigs in different age groups. In this study, we discovered that overexpression of miR-323-3p repressed myoblast proliferation and promoted differentiation, whereas the inhibitor of miR-323-3p displayed the opposite results. Furthermore, we predicted Smad2 as the target gene of miR-323-3p and found that miR-323-3p directly modulated the expression level of Smad2. Then luciferase reporter assays verified that Smad2 was a target gene of miR-323-3p during the differentiation of myoblasts. These findings reveal that miR-323-3p is a positive regulator of myogenesis by targeting Smad2. This provides a novel mechanism of miRNAs in myogenesis.     

Vocal divergence is concordant with genomic evidence for strong reproductive isolation in grasshopper mice (Onychomys)

Behavioral barriers to gene flow often evolve faster than intrinsic incompatibilities and can eliminate the opportunity for hybridization between interfertile species. While acoustic signal divergence is a common driver of premating isolation in birds and insects, its contribution to speciation in mammals is less studied. Here we characterize the incidence of, and potential barriers to, hybridization among three closely related species of grasshopper mice (genus Onychomys). All three species use long‐distance acoustic signals to attract and localize mates; Onychomys arenicola and Onychomys torridus are acoustically similar and morphologically cryptic whereas Onychomys leucogaster is larger and acoustically distinct. We used genotyping‐by‐sequencing (GBS) to test for evidence of introgression in 227 mice from allopatric and sympatric localities in the western United States and northern Mexico. We conducted laboratory mating trials for all species pairs to assess reproductive compatibility, and recorded vocalizations from O. arenicola and O. torridus in sympatry and allopatry to test for evidence of acoustic character displacement. Hybridization was rare in nature and, contrary to prior evidence for O. torridus/O. arenicola hybrids, only involved O. leucogaster and O. arenicola. In contrast, laboratory crosses between O. torridus and O. arenicola produced litters whereas O. leucogaster and O. arenicola crosses did not. Call fundamental frequency in O. torridus and O. arenicola was indistinguishable in allopatry but significantly differentiated in sympatry, a pattern consistent with reproductive character displacement. These results suggest that assortative mating based on a long‐distance signal is an important isolating mechanism between O. torridus and O. arenicola and highlight the importance of behavioral barriers in determining the permeability of species boundaries.     

Modification and evaluation of the Carba NP test by use of paper strip for simple and rapid detection of carbapenemase-producing <Emphasis Type="Italic">Enterobacteriaceae</Emphasis>

Carbapenemase-producing Enterobacteriaceae (CPE) isolates have now emerged worldwide. We therefore modified the phenotypic Carba NP test by use of a filter paper strip for easily and rapidly identifying CPE in routine laboratory. A collection of 56 CPE and carbapenemase-producing Pseudomonas spp. isolates (including 28 NDM-1, 11 IMP-14a, 1 IMP-1, 1 IMP-4, 1 IMP-9, 1 IMP-15, 4 VIM-2, 1 VIM-1, 1 IMP-14a with VIM-2, 3 OXA-48, 3 OXA-181 and 1 KPC-2 producers) and 41 non-CPE isolates (including 19 ESBL, 7 pAmpC, 3 AmpC, 9 ESBL with pAmpC and 3 non-ESBL & non-AmpC producers) as confirmed by the PCR methods were tested by the paper strip method using pharmaceutical imipenem/cilastatin as a substrate. Bacterial colonies of each isolate were applied directly on filter paper strips dropped with either imipenem-phenol red (test strip) or phenol red solution alone (control strip). The reaction was read within 5 min. This test failed to detect 3 OXA-181, 2 OXA-48 and 3 IMP-14a producers (85.7 % sensitivity), whereas no false positives were seen (100 % specificity). Further evaluation of the paper strip test in 267 CPE screening-positive isolates from three hospitals by their medical technologists showed 92.0 % sensitivity (100 % for NDM producers) and 100 % specificity compared with the PCR methods. Because of its ease, rapidness and cost effective, the paper strip test has a potential for routine CPE testing in low-resource laboratories particularly in areas with high prevalence of NDM enzymes, leading to appropriate antimicrobial therapy and infection control strategy.