Oligonucleotide Insecticides for Green Agriculture: Regulatory Role of Contact DNA in Plant–Insect Interactions

Insects vastly outnumber us in terms of species and total biomass, and are among the most efficient and voracious consumers of plants on the planet. As a result, to preserve crops, one of the primary tasks in agriculture has always been the need to control and reduce the number of insect pests. The current use of chemical insecticides leads to the accumulation of xenobiotics in ecosystems and a decreased number of species in those ecosystems, including insects. Sustainable development of human society is impossible without useful insects, so the control of insect pests must be effective and selective at the same time. In this article, we show for the first time a natural way to regulate the number of insect pests based on the use of extracellular double-stranded DNA secreted by the plant Pittosporum tobira. Using a principle similar to one found in nature, we show that the topical application of artificially synthesized short antisense oligonucleotide insecticides (olinscides, DNA insecticides) is an effective and selective way to control the insect Coccus hesperidum. Using contact oligonucleotide insecticide Coccus-11 at a concentration of 100 ng/μL on C. hesperidum larvae resulted in a mortality of 95.59 ± 1.63% within 12 days. Green oligonucleotide insecticides, created by nature and later discovered by humans, demonstrate a new method to control insect pests that is beneficial and safe for macromolecular insect pest management.     

Usnic Acid-Mediated Exchange of Protons for Divalent Metal Cations across Lipid Membranes: Relevance to Mitochondrial Uncoupling

Usnic acid (UA), a unique lichen metabolite, is a protonophoric uncoupler of oxidative phosphorylation, widely known as a weight-loss dietary supplement. In contrast to conventional proton-shuttling mitochondrial uncouplers, UA was found to carry protons across lipid membranes via the induction of an electrogenic proton exchange for calcium or magnesium cations. Here, we evaluated the ability of various divalent metal cations to stimulate a proton transport through both planar and vesicular bilayer lipid membranes by measuring the transmembrane electrical current and fluorescence-detected pH gradient dissipation in pyranine-loaded liposomes, respectively. Thus, we obtained the following selectivity series of calcium, magnesium, zinc, manganese and copper cations: Zn²⁺ > Mn²⁺ > Mg²⁺ > Ca²⁺ >> Cu²⁺. Remarkably, Cu²⁺ appeared to suppress the UA-mediated proton transport in both lipid membrane systems. The data on the divalent metal cation/proton exchange were supported by circular dichroism spectroscopy of UA in the presence of the corresponding cations.     

The Reactions of N,N′-Diphenyldithiomalondiamide with Arylmethylidene Meldrum’s Acids

The Michael addition reaction between dithiomalondianilide (N,N′-diphenyldithiomalondiamide) and arylmethylidene Meldrum’s acids, accompanied by subsequent heterocyclization, was investigated along with factors affecting the mixture composition of the obtained products. The plausible mechanism includes the formation of stable Michael adducts which, under the studied conditions, undergo further transformations to yield corresponding N-methylmorpholinium 4-aryl-6-oxo-3-(N-phenylthio-carbamoyl)-1,4,5,6-tetrahydropyridin-2-thiolates and their oxidation derivatives, 4,5-dihydro-3H-[1,2]dithiolo[3,4-b]pyridin-6(7H)-ones. The structure of one such product, N-methylmorpholinium 2,2-dimethyl-5-(1-(2-nitrophenyl)-3-(phenylamino)-2-(N-phenylthiocarbamoyl)-3-thioxopropyl)-4-oxo-4H-1,3-dioxin-6-olate, was confirmed via X-ray crystallography.     

中国沙棘乙醇提取物的体外及对HepG2细胞的抗氧化活性

本文从生化角度全面分析了中国沙棘乙醇提取物的体外抗氧化活性,并在评价细胞毒性的基础上,以Hep G2细胞为模型,利用流式细胞仪评估了提取物的细胞抗氧化能力。结果表明,中国沙棘乙醇提取物对ABTS自由基具有良好的清除效果,当浓度为5 mg/m L时,其清除能力与阳性对照BHT相当;中国沙棘乙醇提取物对NO自由基也有良好的清除效果,但其整体清除能力不及BHT;当浓度为5 mg/m L时,中国沙棘乙醇提取物对亚油酸脂质过氧化表现抑制,但其抑制效果低于BHT;中国沙棘乙醇提取物的总抗氧化能力随浓度的增大而增强,当浓度为5 mg/m L时,其总抗氧化能力与同浓度下的BHT接近。在浓度为0.05~5 mg/m L范围内,中国沙棘乙醇提取物对Hep G2细胞不显示毒性;当处理浓度为0.5、1、5 mg/m L时,中国沙棘乙醇提取物的阳性细胞率与阳性对照相比分别降低了75.22%、72.36%和78.2%。中国沙棘乙醇提取物不仅在体外条件下具有良好的抗氧化活性,而且在Hep G2细胞内表现出良好的抗氧化能力。      

Physiologic Insulin Resensitization as a Treatment Modality for Insulin Resistance Pathophysiology

Prevalence of type 2 diabetes increased from 2.5% of the US population in 1990 to 10.5% in 2018. This creates a major public health problem, due to increases in long-term complications of diabetes, including neuropathy, retinopathy, nephropathy, skin ulcers, amputations, and atherosclerotic cardiovascular disease. In this review, we evaluated the scientific basis that supports the use of physiologic insulin resensitization. Insulin resistance is the primary cause of type 2 diabetes. Insulin resistance leads to increasing insulin secretion, leading to beta-cell exhaustion or burnout. This triggers a cascade leading to islet cell destruction and the long-term complications of type 2 diabetes. Concurrent with insulin resistance, the regular bursts of insulin from the pancreas become irregular. This has been treated by the precise administration of insulin more physiologically. There is consistent evidence that this treatment modality can reverse the diabetes-associated complications of neuropathy, diabetic ulcers, nephropathy, and retinopathy, and that it lowers HbA1c. In conclusion, physiologic insulin resensitization has a persuasive scientific basis, significant treatment potential, and likely cost benefits.     

Immature Vascular Smooth Muscle Cells in Healthy Murine Arteries and Atherosclerotic Plaques: Localization and Activity

The local development of atherosclerotic lesions may, at least partly, be associated with the specific cellular composition of atherosclerosis-prone regions. Previously, it was demonstrated that a small population of immature vascular smooth muscle cells (VSMCs) expressing both CD146 and neuron-glial antigen 2 is postnatally sustained in atherosclerosis-prone sites. We supposed that these cells may be involved in atherogenesis and can continuously respond to angiotensin II, which is an atherogenic factor. Using immunohistochemistry, flow cytometry, wound migration assay xCELLigence system, and calcium imaging, we studied the functional activities of immature VSMCs in vitro and in vivo. According to our data, these cells do not express nestin, CD105, and the leptin receptor. They are localized in atherosclerosis-prone regions, and their number increases with age, from 5.7% to 23%. Immature VSMCs do not migrate to low shear stress areas and atherosclerotic lesions. They also do not have any unique response to angiotensin II. Thus, despite the localization of immature VSMCs and the presence of the link between their number and age, our study did not support the hypothesis that immature VSMCs are directly involved in the formation of atherosclerotic lesions. Additional lineage tracing studies can clarify the fate of these cells during atherogenesis.     

Local Accumulation of Lymphocytes in the Intima of Human Aorta Is Associated with Giant Multinucleated Endothelial Cells: Possible Explanation for Mosaicism of Atherosclerosis

Distribution of different types of atherosclerotic lesions in the arterial wall is not diffuse, but is characterized by mosaicism. The causes of such distribution remain to be established. At the early stages of atherogenesis, low-density lipoprotein (LDL) particles and immune cells penetrate into the intimal layer of the arterial wall through the endothelium. In adult humans, the luminal surface of the arterial wall is a heterogeneous monolayer of cells with varying morphology including typical endothelial cells (ECs) and multinucleated variant endothelial cells (MVECs). We hypothesized that distribution of MVECs in the endothelial monolayer can be related to the distribution pattern of early atherosclerotic lesions. We obtained en face preparations of intact adult (22–59 years old) aortic wall sections that allowed us to study the endothelial monolayer and the subendothelial layer. We compared the distribution of MVECs in the endothelial monolayer with the localization of early atherosclerotic lesions in the subendothelial layer, which were characterized by lipid accumulation and immune cell recruitment. In primary culture, MVECs demonstrated increased phagocytic activity compared to mononuclear ECs. Moreover, we have shown that unaffected aortic intima contained associates formed as a result of aggregation and/or fusion of LDL particles that are non-randomly distributed. This indicated that MVECs may be involved in the accumulation of LDL in the subendothelial layer through increased transcytosis. Interaction of LDL with subendothelial cells of human aorta in primary culture increased their adhesive properties toward circulating immune cells. Study of unaffected aortic intima revealed non-random distribution of leukocytes in the subendothelial layer and increased localization of CD45+ leukocytes in the subendothelial layer adjacent to MVECs. Together, our observations indicate that MVECs may be responsible for the distribution of atherosclerotic lesions in the arterial wall by participating in LDL internalization and immune cell recruitment.     

Profiling of the Bacterial Microbiota along the Murine Alimentary Tract

Here, the spatial distribution of the bacterial flora along the murine alimentary tract was evaluated using high throughput sequencing in wild-type and Tff3-deficient (Tff3^KO) animals. Loss of Tff3 was linked to increased dextran sodium sulfate-induced colitis. This systematic study shows the results of 13 different regions from the esophagus to the rectum. The number of bacterial species (richness) increased from the esophagus to the rectum, from 50 to 200, respectively. Additionally, the bacterial community structure changed continuously; the highest changes were between the upper/middle and lower gastrointestinal compartments when comparing adjacent regions. Lactobacillus was the major colonizer in the upper/middle gastrointestinal tract, especially in the esophagus and stomach. From the caecum, a drastic diminution of Lactobacillus occurred, while members of Lachnospiraceae significantly increased. A significant change occurred in the bacterial community between the ascending and the transverse colon with Bacteroidetes being the major colonizers with relative constant abundance until the rectum. Interestingly, wild-type and Tff3^KO animals did not show significant differences in their bacterial communities, suggesting that Tff3 is not involved in alterations of intraluminal or adhesive microbiota but is obviously important for mucosal protection, e.g., of the sensitive stem cells in the colonic crypts probably by a mucus plume.     

Nutritional Calcium Supply Dependent Calcium Balance,Bone Calcification and Calcium Isotope Ratios in Rats

Serum calcium isotopes (δ^44/42Ca) have been suggested as a non-invasive and sensitive Ca balance marker. Quantitative δ^44/42Ca changes associated with Ca flux across body compartment barriers relative to the dietary Ca and the correlation of δ^44/42Ca_Serum with bone histology are unknown. We analyzed Ca and δ^44/42Ca by mass-spectrometry in rats after two weeks of standard-Ca-diet (0.5%) and after four subsequent weeks of standard- and of low-Ca-diet (0.25%). In animals on a low-Ca-diet net Ca gain was 61 ± 3% and femur Ca content 68 ± 41% of standard-Ca-diet, bone mineralized area per section area was 68 ± 15% compared to standard-Ca-diet. δ^44/42Ca was similar in the diets, and decreased in feces and urine and increased in serum in animals on low-Ca-diet. δ^44/42Ca_Bone was higher in animals on low-Ca-diet, lower in the diaphysis than the metaphysis and epiphysis, and unaffected by gender. Independent of diet, δ^44/42Ca_Bone was similar in the femora and ribs. At the time of sacrifice, δ^44/42Ca_Serum inversely correlated with intestinal Ca uptake and histological bone mineralization markers, but not with Ca content and bone mineral density by µCT. In conclusion, δ^44/42Ca_Bone was bone site specific, but mechanical stress and gender independent. Low-Ca-diet induced marked changes in feces, serum and urine δ^44/42Ca in growing rats. δ^44/42Ca_Serum inversely correlated with markers of bone mineralization.