Therapeutic efficacy of Stephania tetrandra S. Moore for treatment of neovascularization of retinal capillary (retinopathy) in diabetes--in vitro study.

The present study was designed to examine therapeutic efficacy of the root extract of Stephania Tetrandra S. Moore (STMS) (traditional Chinese medicine; Han Fang Ji) for treatment of neovascularization of the retinal capillary (retinopathy) in streptozotocin (STZ)-induced diabetic rats (STZ diabetic rats) in culture. Recently we have established the culture system in which fetal bovine serum (FBS) in Dulbecco modified Eagle medium (DMEM) induced neovascularization of the retinal capillary and choroidal capillary in normal rats in culture. STZ diabetic rats showed more neovascularization of the retinal capillary and choroidal capillary than did normal rats in culture. In this study, the retinal tissue was removed for the posterior ocular region and cultured in DMEM containing FBS. The choroidal tissue of the posterior ocular region was also removed and cultured as an internal reference. Administration of STSM (0.91, 9.1 and 91 microg/ml) significantly suppressed neovascularization of the retinal capillary in both STZ diabetic rats and normal rats in a dose-dependent manner. Similar results were obtained with the choroidal capillary; administration of STSM suppressed neovascularization of the choroidal capillary in both STZ diabetic rats and normal rats. In order to determine the component of STSM inhibiting neovascularization of the retinal capillary, tetrandrine (a major chemical constituent of STSM) was administered and neovascularization of the retinal capillary was examined in culture. The effect of tetrandrine on the choroidal capillary was also examined as an internal reference. Administration of tetrandrine (0.1, 1.0 and 10 microM) suppressed neovascularization of the retinal capillary in both STZ diabetic rats and normal rats in a dose-dependent manner. Similar results were obtained with the choroidal capillary of both STZ diabetic rats and normal rats. We infer, therefore, that STSM has a direct effect on the retinal capillary of posterior ocular region and suppresses neovascularization of retinal capillary in STZ diabetic rats through the activation of tetrandrine. These results suggest that STSM may prevent for delay the progression of retinopathy in diabetic patients.     

Active compounds in Populus nigra L. wilted leaves responsible for attracting Helicoverpa armigera (Hübner) (Lep., Noctuidae) and new agaropectin formulation

Abstract:  The leaf extracts of Populus nigra were collected and identified by steam distillation, air entrainment and gas chromatographic–mass spectrometric analysis. Electroantennograms were recorded from Helicoverpa armigera adults in response to the chemicals identified. Both aromatic compounds and green-leaf volatiles elicited strong responses. Field experiments revealed that the active compounds responsible for attracting H. armigera moths are mainly short-side-chain aromatic alcohols and aldehydes. We, for the first time, used agaropectin as the controlled-release matrix of insect attractants. A five-component lure containing all the aromatics without phenolics, mixed in the proportions as found in the steam distillate of the leaves collected in August, produced the best trap catch. The results showed that the volatiles of wilted leaves of P. nigra can attract H. armigera adults by feeding attraction.     

Pre‐copulation intervals,copulation frequencies,and initial progeny sex ratios in two biotypes of whitefly,Bemisia tabaci

The whitefly Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae) is a species complex, and its systematic classification requires controlled crossing experiments among its genetic groups. Accurate information on pre‐copulation intervals, copulation frequencies, and initial frequency of egg fertilization of newly emerged adults is critical for designing procedures for collecting the virgin adults necessary for these experiments. In the literature, considerable variation is reported between B. tabaci populations, with respect to the length of the pre‐copulation interval and the initial frequency of egg fertilization. Here, we used a video‐recording method to observe continuously the copulation behaviour of the Mediterranean/Asia Minor/Africa (B biotype) and the Asia II (ZHJ1 biotype) groups of B. tabaci. We also recorded the initial frequency of egg fertilization, as determined by the sex of the progeny. When adults were caged in female–male pairs on leaves of cotton plants, the earliest copulation events occurred 2–6 h after emergence; at 12 h after emergence 56–84% of the females had copulated at least once, and nearly all (92–100%) had copulated at least once by 36 h after emergence. Both females and males copulated repeatedly. Approximately 80 and 20% of copulation events occurred during the photophase and scotophase, respectively. By 72 h post‐emergence, the females of the B and ZHJ1 biotypes had copulated on average 6.1 and 3.9 times, respectively. When adults were caged in groups on plants 1–13 h after emergence, 30–35% of the eggs deposited during this period were fertilized, and approximately 90% of females were fertilized by the end of the 13 h. Although timing of copulation differed in detail between the two genetic groups, the results demonstrate that B. tabaci adults can start to copulate as early as 2–6 h post‐emergence and the majority of females can become fertilized on the day that they emerge.     

EFFECTS OF HELPER PROTEINS ENCODED BY p19 AND orf1-orf2 GENES ON Cyt1Aa PROTEIN EXPRESSION IN ACRYSTALLIFEROUS STRAIN OF BACILLUS THURINGIENSIS

A series of plasmids were constructed to examine the effects of p19 and orf1‐orf2 genes from Bacillus thuringiensis on Cyt1Aa synthesis and inclusion formation. The plasmids expressed the cyt1Aa gene along with either p19 or orf1‐orf2, or each of them coordinatively with p20 in the acrystalliferous strain of B. thuringiensis subsp. israelensis 4Q7. No effect on the expression of Cyt1Aa protein was found when P19 or Orf1‐Orf2 co‐expressed with Cyt1Aa. However, when including p20 gene, the constructs with p19 or orf1‐orf2 gene produced lower yield of Cyt1Aa proteins than without p19 or orf1‐orf2 gene. Electron microscopy observation and bioassay showed that P19 and Orf1‐Orf2 have no influence on the crystal size and toxicity of Cyt1Aa protein. It is presumed that P19 and Orf1‐Orf2 might have negative effects on Cyt1Aa synthesis in B. thuringiensis.     

RFLP tagging of a salt tolerance gene in rice

A salt tolerant rice mutant (M-20) was obtained through selection in vitro. Its tolerance was stably inherited over eight generations and most traints between M-20 and its sensitive original 77–170 (Oryza sativa) were very similar. By deriving an F₂ population of M-20 × 77–170 and splitting every F₂ individual into two parts, with one part planted in normal conditions and another part in saline conditions, the inheritance of salt tolerance in rice was studied. Under normal conditions, there was no apparent segregation among F₂ individuals. Under saline conditions, however, the segregation of traits was obvious. According to our standards, the ratio of salt sensitive:moderately-tolerant:tolerant plants was 25:42:18, in accordance with a 1:2:1 ratio. It suggested that the improvement of salt tolerance in our materials was induced by the mutation of a major tolerant gene which showed incomplete dominance. By use of 130 RFLP probes distributed throughout the rice genome, the gene was tagged by a single copy DNA probe, RG4, which was located on chromosome 7. The genetic distance between the salt tolerant gene and RG4 was 7.0 ± 2.9 cM. Based on the split method, a method which could be currently used to evaluate the damage of salt stress in rice was proposed.     

Adenine affects the structure and stability of telomeric sequences.

Adenine occurs in the strand containing repeated G clusters in the telomeric DNA of a variety of organisms, including that of humans. The role of adenine has been investigated by constructing two sets of oligonucleotides each with one, two, or four copies of the telomeric sequence dTTTAGGG together with a control sequence in which T replaces the A residue, dTTTTGGG. Comparison of the stability and spectral properties of these two sequences in the presence of Na+ or K+ affords a basis for defining the role of adenine in these structures. In Na+, the A residue stabilizes the structure formed by each oligomer significantly, presumably by a base-pairing interaction with T. In K+, by contrast, there is little difference in stability. In two- and four-copy oligomers, the A sequence has a different structure from its T analog, as detected by CD spectroscopy. In the presence of either Na+ or K+, the tetraplexes of A and T interact with intercalators.     

Chromosome-level genome assembly and characterization of Sophora Japonica

Sophora japonica is a medium-size deciduous tree belonging to Leguminosae family and famous for its high ecological, economic and medicinal value. Here, we reveal a draft genome of S. japonica, which was ∼511.49 Mb long (contig N50 size of 17.34 Mb) based on Illumina, Nanopore and Hi-C data. We reliably assembled 110 contigs into 14 chromosomes, representing 91.62% of the total genome, with an improved N50 size of 31.32 Mb based on Hi-C data. Further investigation identified 271.76 Mb (53.13%) of repetitive sequences and 31,000 protein-coding genes, of which 30,721 (99.1%) were functionally annotated. Phylogenetic analysis indicates that S. japonica separated from Arabidopsis thaliana and Glycine max ∼107.53 and 61.24 million years ago, respectively. We detected evidence of species-specific and common-legume whole-genome duplication events in S. japonica. We further found that multiple TF families (e.g. BBX and PAL) have expanded in S. japonica, which might have led to its enhanced tolerance to abiotic stress. In addition, S. japonica harbours more genes involved in the lignin and cellulose biosynthesis pathways than the other two species. Finally, population genomic analyses revealed no obvious differentiation among geographical groups and the effective population size continuously declined since 2 Ma. Our genomic data provide a powerful comparative framework to study the adaptation, evolution and active ingredients biosynthesis in S. japonica. More importantly, our high-quality S. japonica genome is important for elucidating the biosynthesis of its main bioactive components, and improving its production and/or processing.     

SARS-CoV-2 envelope protein causes acute respiratory distress syndrome (ARDS)-like pathological damages and constitutes an antiviral target

Cytokine storm and multi-organ failure are the main causes of SARS-CoV-2-related death. However, the origin of excessive damages caused by SARS-CoV-2 remains largely unknown. Here we show that the SARS-CoV-2 envelope (2-E) protein alone is able to cause acute respiratory distress syndrome (ARDS)-like damages in vitro and in vivo. 2-E proteins were found to form a type of pH-sensitive cation channels in bilayer lipid membranes. As observed in SARS-CoV-2-infected cells, heterologous expression of 2-E channels induced rapid cell death in various susceptible cell types and robust secretion of cytokines and chemokines in macrophages. Intravenous administration of purified 2-E protein into mice caused ARDS-like pathological damages in lung and spleen. A dominant negative mutation lowering 2-E channel activity attenuated cell death and SARS-CoV-2 production. Newly identified channel inhibitors exhibited potent anti-SARS-CoV-2 activity and excellent cell protective activity in vitro and these activities were positively correlated with inhibition of 2-E channel. Importantly, prophylactic and therapeutic administration of the channel inhibitor effectively reduced both the viral load and secretion of inflammation cytokines in lungs of SARS-CoV-2-infected transgenic mice expressing human angiotensin-converting enzyme 2 (hACE-2). Our study supports that 2-E is a promising drug target against SARS-CoV-2.Subject terms: Cell death, Molecular biology     

Participation of lipopolysaccharide in hyperplasic adipose expansion: Involvement of NADPH oxidase/ROS/p42/p44 MAPK‐dependent Cyclooxygenase‐2

Obesity is a world‐wide problem, especially the child obesity, with the complication of various metabolic diseases. Child obesity can be developed as early as the age between 2 and 6. The expansion of fat mass in child age includes both hyperplasia and hypertrophy of adipose tissue, suggesting the importance of proliferation and adipogenesis of preadipocytes. The changed composition of gut microbiota is associated with obesity, revealing the roles of lipopolysaccharide (LPS) on manipulating adipose tissue development. Studies suggest that LPS enters the circulation and acts as a pro‐inflammatory regulator to facilitate pathologies. Nevertheless, the underlying mechanisms behind LPS‐modulated obesity are yet clearly elucidated. This study showed that LPS enhanced the expression of cyclooxygenase‐2 (COX‐2), an inflammatory regulator of obesity, in preadipocytes. Pretreating preadipocytes with the scavenger of reactive oxygen species (ROS) or the inhibitors of NADPH oxidase or p42/p44 MAPK markedly decreased LPS‐stimulated gene expression of COX‐2 together with the phosphorylation of p47^phox and p42/p44 MAPK, separately. LPS activated p42/p44 MAPK via NADPH oxidase‐dependent ROS accumulation in preadipocytes. Reduction of intracellular ROS or attenuation of p42/p44 MAPK activation both reduced LPS‐mediated COX‐2 expression and preadipocyte proliferation. Moreover, LPS‐induced preadipocyte proliferation and adipogenesis were abolished by the inhibition of COX‐2 or PEG₂ receptors. Taken together, our results suggested that LPS enhanced the proliferation and adipogenesis of preadipocytes via NADPH oxidase/ROS/p42/p44 MAPK‐dependent COX‐2 expression.