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Titanium nanoparticles and pyrethroid pesticides are now being widely used in industrial, agriculture, and biomedical applications. In recent years, their health safety profiles have aroused concerns among health scientists. This study mainly explored the combined effects of titanium dioxide nanoparticles (TiO

Introduction Transcription factors (TFs) and cis-regulatory elements (CREs) control gene transcripts involved in various biological processes. We hypothesize that TFs and CREs can be effective molecular tools for De Novo regulation designs to engineer plants. Objectives We selected two Arabidopsis TF types and two tobacco CRE types to design a De Novo regulation and evaluated its effectiveness in plant engineering. Methods G-box and MYB recognition elements (MREs) were identified in four Nicotiana tabacum JAZs (NtJAZs) promoters. MRE-like and G-box like elements were identified in one nicotine pathway gene promoter. TF screening led to select Arabidopsis Production of Anthocyanin Pigment 1 (PAP1/MYB) and Transparent Testa 8 (TT8/bHLH). Two NtJAZ and two nicotine pathway gene promoters were cloned from commercial Narrow Leaf Madole (NL) and KY171 (KY) tobacco cultivars. Electrophoretic mobility shift assay (EMSA), cross-linked chromatin immunoprecipitation (ChIP), and dual-luciferase assays were performed to test the promoter binding and activation by PAP1 (P), TT8 (T), PAP1/TT8 together, and the PAP1/TT8/Transparent Testa Glabra 1 (TTG1) complex. A DNA cassette was designed and then synthesized for stacking and expressing PAP1 and TT8 together. Three years of field trials were performed by following industrial and GMO protocols. Gene expression and metabolic profiling were completed to characterize plant secondary metabolism. Results PAP1, TT8, PAP1/TT8, and the PAP1/TT8/TTG1 complex bound to and activated NtJAZ promoters but did not bind to nicotine pathway gene promoters. The engineered red P + T plants significantly upregulated four NtJAZs but downregulated the tobacco alkaloid biosynthesis. Field trials showed significant reduction of five tobacco alkaloids and four carcinogenic tobacco specific nitrosamines in most or all cured leaves of engineered P + T and PAP1 genotypes. Conclusion G-boxes, MREs, and two TF types are appropriate molecular tools for a De Novo regulation design to create a novel distant-pathway cross regulation for altering plant secondary metabolism.

Titanium dioxide nanoparticles (TiO2NPs) and cypermethrin (CPM) are widely used in various fields, and they can enter the environment in different ways. Combined exposure of TiO2NPs and CPM may increase the accumulation of pollutants in organisms and affect human health. This study was undertaken to evaluate the oxidative and inflammatory parameters associated with the combined exposure of TiO2NPs and CPM in rats. Twenty-four healthy male adult SD rats were randomly divided into four groups. The first group served as the control, while groups 2, 3, and 4 were treated with TiO2NPs (450 mg/m3); CPM (6.67 mg/m3) or combined exposure of TiO2NPs and CPM by inhalation for 90 days. We investigated the oxidative damage induced through combined exposure of TiO2NPs and CPM in rats by evaluating hematology of the rats and determining the blood biochemical index. Our results demonstrated that inhalation of TiO2NPs and CPM increased the levels of oxidative stress markers such as malondialdehyde and alkaline phosphatase in the serum of rats. These were accompanied by a decreased glutathione peroxidase and total superoxide dismutase levels. Furthermore, the level of glutathione peroxidase was further decreased while malondialdehyde was increased in the combined exposure of TiO2NPs and CPM. Interestingly, pathological sections showed that different degrees of tissue injury could be seen in the liver and lung tissues of each exposure group. In summary, the combined exposure of TiO2NPs and CPM can cause increased oxidative damage in rats and damage the tissue structure of the liver and lung.

Objective: To explore the optimal ablation index (AI) parameters for radiofrequency catheter ablation (RA) for treating atrial fibrillation (AF). Method: Patients with AF (186) who underwent bilateral PVAI in the Department of Cardiology, Zhuhai People’s Hospital, Guangdong Province, from March 2018 to October 2019 and received catheter ablation as first-round treatment, were grouped according to the received AI. Control group included patients (95) who received the recommended AI ablation (350–400 for posterior wall, 400–450 for non-posterior wall). Patients in optimal AI group were ablated with optimal AI (300–330 for posterior wall, 350–380 for non-posterior wall). Results: Of 186 patients, 66 patients had paroxysmal atrial fibrillation and a mean CHA2DS2-VASc score of 2.83±1.64. Isolation rates of bilateral PVI in both groups were 91.4% and 93.6%, for patients with paroxysmal atrial fibrillation, and 81.7% and 80% for patients with persistent atrial fibrillation (P > 0.05). Left atrial function index (LAFI) decreased under the condition of sinus rhythm at the 3rd and 6th months (P < 0.05). LAFI improvement was significantly better in the optimal AI group than in the control group (P < 0.05). Rates of pain and cough during the ablation, and postoperative gastrointestinal discomfort and use of PPIs were higher in the control group (P < 0.05). Conclusion: Radiofrequency ablation of AF, guided by optimal AI combined with impedance, can minimize atrial injury, prevent atrial failure, promote the recovery of atrial function, reduces intraoperative cough, pain, and postoperative gastrointestinal discomfort and use of PPIs. doi: https://doi.org/10.12669/pjms.38.3.4971 How to cite this:Qin X, Jiang X, Yuan Q, Xu G, He X. Optimal ablation index parameters for radiofrequency ablation therapy of atrial fibrillation. Pak J Med Sci. 2022;38(3):---------. doi: https://doi.org/10.12669/pjms.38.3.4971 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Seasonally frozen soil where uneven freeze–thaw damage is a major cause of highway deterioration has attracted increased attention in China with the rapid development of infrastructure projects. Based on Darcy’s law of unsaturated soil seepage and heat conduction, the thermal–hydraulic–mechanical (THM) coupling model is established considering a variety of effects (i.e., ice–water phase transition, convective heat transfer, and ice blocking effect), and then the numerical solution of thermal–hydraulic fields of subgrade can be obtained. Then, a new concept, namely degree of freeze–thaw damage, is proposed by using the standard deviation of the ice content of subgrade during the annual freeze–thaw cycle. To analyze the freeze–thaw characteristics of highway subgrade, the model is applied in the monitored section of the Golmud to Nagqu portion of China National Highway G109. The results show that: (1) The hydrothermal field of subgrade has an obvious sunny–shady slopes effect, and its transverse distribution is not symmetrical, (2) the freeze–thaw damage area of subgrade obviously decreased under the insulation board measure, (3) under the combined anti-frost measures, the maximum frost heave amount of subgrade is significantly reduced. This study will provide references for the design of highway subgrades in seasonally frozen soil areas.

Transcription factors (TFs) andWe selected two Arabidopsis TF types and two tobacco CRE types to design aG-box and MYB recognition elements (MREs) were identified in fourPAP1, TT8, PAP1/TT8, and the PAP1/TT8/TTG1 complex bound to and activatedG-boxes, MREs, and two TF types are appropriate molecular tools for a

IntroductionTranscription factors (TFs) and cis-regulatory elements (CREs) control gene transcripts involved in various biological processes. We hypothesize that TFs and CREs can be effective molecular tools for De Novo regulation designs to engineer plants.ObjectivesWe selected two Arabidopsis TF types and two tobacco CRE types to design a De Novo regulation and evaluated its effectiveness in plant engineering.MethodsG-box and MYB recognition elements (MREs) were identified in four Nicotiana tabacum JAZs (NtJAZs) promoters. MRE-like and G-box like elements were identified in one nicotine pathway gene promoter. TF screening led to select Arabidopsis Production of Anthocyanin Pigment 1 (PAP1/MYB) and Transparent Testa 8 (TT8/bHLH). Two NtJAZ and two nicotine pathway gene promoters were cloned from commercial Narrow Leaf Madole (NL) and KY171 (KY) tobacco cultivars. Electrophoretic mobility shift assay (EMSA), cross-linked chromatin immunoprecipitation (ChIP), and dual luciferase assays were performed to test the promoter binding and activation by PAP1 (P), TT8 (T), PAP1/TT8 together, and the PAP1/TT8/Transparent Testa Glabra 1 (TTG1) complex. A DNA cassette was designed and then synthesized for stacking and expressing PAP1 and TT8 together. Three years of field trials were performed by following industrial and GMO protocols. Gene expression and metabolic profiling were completed to characterize plant secondary metabolism.ResultsPAP1, TT8, PAP1/TT8, and the PAP1/TT8/TTG1 complex bound to and activated NtJAZ promoters but did not bind to nicotine pathway gene promoters. The engineered red P+T plants significantly upregulated four NtJAZs but downregulated the tobacco alkaloid biosynthesis. Field trials showed significant reduction of five tobacco alkaloids and four carcinogenic tobacco specific nitrosamines in most or all cured leaves of engineered P+T and PAP1 genotypes.ConclusionG-boxes, MREs, and two TF types are appropriate molecular tools for a De Novo regulation design to create a novel distant-pathway cross regulation for altering plant secondary metabolism.

Objective: To explore the effects of alternate-day-fasting (ADF) therapy combined with Lingguizhugan Decoction (LD) on blood lipid profiles of hyperlipidemic rats. Methods: Rats were randomly assigned into high-fat-diet (HF) group and normal-diet (ND) group. Hyperlipidemic rats fed with high-fat-diet for 5 weeks were randomly divided into ADF group, alternate-day-fasting with LD (ALG) group and model control (MC) group. The rats in ALG and ADF group were deprived of food for 24 h every other day for 4 weeks. Rats in ALG group were administrated with LD at fasting day. After 4 weeks of ADF therapy, plasma TC, TG, LDL-c and HDL-c were measured in each group. Expression of miR-143 and PPAR-γ protein from adipose was also analyzed. Results: When compared with MC group, after 4 weeks of ADF or combined ADF and LD therapy, the body weight was evidently reduced in ADF and ALG groups (P = 0.028, P = 0.036 by wk 8). The levels of plasma TC and TG decreased in ADF group and ALG group, which were significantly lower than those in MC group (P < 0.001, P = 0.045; P < 0.001, P = 0.005). However, the body weight and level of TC and TG in ALG group showed non-statistical difference in comparison with ADF group (ALG vs. ADF, P > 0.05). Expression of miR-143 and PPAR-γ were higher in MC group than that in NC group (P < 0.001). Compared with MC group, expression of miR-143 and PPAR-γ were significantly decreased in ADF (P = 0.038, P = 0.015) and ALG (P = 0.007, P < 0.001) groups. When compared with ADF group, expression of miR-143 and PPAR-γ were significantly decreased in ALG (P = 0.041, P = 0.046) group. Conclusion: ADF therapy alone not only reduced blood lipids, but also inhibited miR-143 and PPAR-γ protein expression in visceral adipose tissue. However, LD couldn’t reduce the levels of blood lipid profiles more effectively than using ADF alone. Perhaps the effects of LD combined with ADF in the prevention of hyperlipidemia need further exploration.

In this study, we investigate the metabolic engineering of anthocyanins in two dark tobacco crops (Narrow Leaf Madole and KY171) and evaluate the effects on physiological features of plant photosynthesis. Arabidopsis PAP1 (production of anthocyanin pigment 1) gene (AtPAP1) encodes a R2R3-type MYB transcript factor that is a master component of regulatory complexes controlling anthocyanin biosynthesis. AtPAP1 was introduced to Narrow Leaf Madole and KY171 plants. Multiple transgenic plants developed red/purple pigmentation in different tissues. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that the expression levels of six pathway genes were increased two- to eight-fold in AtPAP1 transgenic plants compared with vector control plants. Dihydroflavonol reductase and anthocyanidin synthase genes that were not expressed in wild-type plants were activated. Spectrophotometric measurement showed that the amount of anthocyanins in AtPAP1 transgenic plants were 400-800 µg g-1 fresh weight (FW). High-performance liquid chromatography (HPLC) analysis showed that one main anthocyanin molecule accounted for approximately 98% of the total anthocyanins. Tandem MS/MS analysis using HPLC coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry identified the main anthocyanin as cyanidin 3-O-rutinoside, an important medicinal anthocyanin. Analysis of photosynthesis rate, chlorophylls and carotenoids contents showed no differences between red/purple transgenic and control plants, indicating that this metabolic engineering did not alter photosynthetic physiological traits. This study shows that AtPAP1 is of significance for metabolic engineering of anthocyanins in crop plants for value-added traits.

Staphylococcal enterotoxins (SEs) produced by Staphylococcus aureus have increasingly given rise to human health and food safety. Genetically engineered small molecular antibody is a useful tool in immuno-detection and treatment for clinical illness caused by SEs. In this study, we constructed the V(L)-V(H) tail-parallel genetically engineered antibody against SEs by using the repertoire of rearranged germ-line immunoglobulin variable region genes. Total RNA were extracted from six hybridoma cell lines that stably express anti-SEs antibodies. The variable region genes of light chain (V(L)) and heavy chain (V(H)) were cloned by reverse transcription PCR, and their classical murine antibody structure and functional V(D)J gene rearrangement were analyzed. To construct the eukaryotic V(H)-V(L) tail-parallel co-expression vectors based on the "5'-V(H)-ivs-IRES-V(L)-3'" mode, the ivs-IRES fragment and V(L) genes were spliced by two-step overlap extension PCR, and then, the recombined gene fragment and V(H) genes were inserted into the pcDNA3.1(+) expression vector sequentially. And then the constructed eukaryotic expression clones termed as p2C2HILO and p5C12HILO were transfected into baby hamster kidney 21 cell line, respectively. Two clonal cell lines stably expressing V(L)-V(H) tail-parallel antibodies against SEs were obtained, and the antibodies that expressed intracytoplasma were evaluated by enzyme-linked immunosorbent assay, immunofluorescence assay, and flow cytometry. SEs can stimulate the expression of some chemokines and chemokine receptors in porcine IPEC-J2 cells; mRNA transcription level of four chemokines and chemokine receptors can be blocked by the recombinant SE antibody prepared in this study. Our results showed that it is possible to get functional V(L)-V(H) tail-parallel genetically engineered antibodies in same vector using eukaryotic expression system.

Porcine bocavirus was first discovered in Swedish pigs with post-weaning multisystemic wasting syndrome (PMWS) in 2009. Many efforts have been implemented to investigate the porcine bocavirus, but it remains enigmatic. In the current study, we utilized data from both China and the USA. The China-derived data included 403 pig samples collected from five provinces, 122 gene sequences from the GenBank database, and 637 old porcine bocavirus (PBoV) cases. The USA-derived data comprised 181 pig samples from 18 states, 39 new gene sequences, and 85 new emerging cases. First, we executed a comprehensive analysis of the disease's prevalence, phylogenetics, evolutionary distances, mutation network, geographical distribution, occurrence frequency, and phylogeographical estimation in both China and the USA. The results showed that the positive rates of PBoV (42.0%, 76/181) in American samples were significantly higher than those (11.4%, 46/403) in the Chinese samples. All PBoV cases from these countries can be divided into six groups: PBoV1 (group 1), PBoV2 (group 2), PBoV3C (group 3), PBoV5 (group 4), PBoV3/4 (group 5), and PBoV6V7V (group 6). PBoV1 and PBoV2 were epidemic strains from 2006 to 2011 in China, whereas the PBoV3 subtypes were epidemic from 2010 to 2012 in China and the USA. At present, PBoV3C (group 3), PBoV5 (group 4), and PBoV3/4 (group 5) are epidemic viruses and co-exist in China and the USA. The geographical distribution of PBoV mainly lies in the east and south coastal areas of China and the central states of the USA. Jiangsu Province and the state of Minnesota were the centers of high occurrence frequency of PBoV with six outbreaks. The old PBoV cases involved 14 provinces and regions of China and North Carolina in the USA, whereas the new emerging cases involved five provinces in China and 13 states in the USA, of which two provinces and 12 states reported for the first time that piglets were infected by PBoV. Hong Kong, Hebei, and Jiangsu Provinces and the states of Minnesota and North Carolina were possibly geographical origins of PBoV in China and America, respectively. These data can help us systematically understand porcine bocavirus in China and America and find effective strategies for its treatment.

The 100 trillion microbes in human gut coevolve with the host and exert significant influences on human health. The gut microbial composition presents dynamic changes correlated with various factors including host genotypes, age, and external environment. Effective manipulation of the gut microbiota through diets (both long-term and short-term diet patterns), probiotics and/or prebiotics, and antibiotics has been proved being potential to prevent from metabolic disorders such as obesity in many studies. The dietary regulation exerts influences on microbial metabolism and host immune functions through several pathways, of which may include selectively bacterial fermentation of nutrients, lower intestinal barrier function, overexpression of genes associated with disorders, and disruptions to both innate and adaptive immunity. Discoveries in the interrelationship between diet, intestinal microbiome, and body immune system provide us novel perceptions to the specific action mechanisms and will promote the development of therapeutic approaches for obesity.

The 100 trillion microbes in human gut coevolve with the host and exert significant influences on human health. The gut microbial composition presents dynamic changes correlated with various factors including host genotypes, age, and external environment. Effective manipulation of the gut microbiota through diets (both long-term and short-term diet patterns), probiotics and/or prebiotics, and antibiotics has been proved being potential to prevent from metabolic disorders such as obesity in many studies. The dietary regulation exerts influences on microbial metabolism and host immune functions through several pathways, of which may include selectively bacterial fermentation of nutrients, lower intestinal barrier function, overexpression of genes associated with disorders, and disruptions to both innate and adaptive immunity. Discoveries in the interrelationship between diet, intestinal microbiome, and body immune system provide us novel perceptions to the specific action mechanisms and will promote the development of therapeutic approaches for obesity.