Construction and validation of co-expression vector for rice alpha tubulin and microtubule associated protein respectively fused with fluorescent proteins.

Microtubule (MT) consists of α-tubulin and ß-tubulin. The dynamic instability regulated by various microtubule associated proteins (MAPs) is essential for MT functions. To analyze the interaction between tubulin/MT and MAP in vivo, we usually need tubulin and MAP co-expressed. Here, we constructed a dual-transgene vector expressing rice (Oryza sativa) α-tubulin and MAP simultaneously. To construct this vector, plant expression vector pCambia1301 was used as the plasmid backbone and Gibson assembly cloning technology was used. We first fused and cloned the GFP fragment, α-tubulin open reading frame (ORF), and NOS terminator into the vector pCambia1301 to construct the p35S::GFP-α-tubulin vector that expressed GFP-α-tubulin fusion protein. Subsequently, we fused and cloned the CaMV 35S promoter, mCherry fragment, and NOS terminator into the p35S::GFP-α-tubulin vector to generate the universal dual-transgene expression vector (p35S::GFP-α-tubulin-p35S::mCherry vector). With the p35S::GFP-α-tubulin-p35S::mCherry vector, MAP ORF can be cloned into the site of 5' or 3' terminus of mCherry to co-express GFP-α-tubulin and MAP-mCherry/mCherry-MAP. To validate the availability and universality of the dual-transgene expression vector, a series of putative rice MAP genes including GL7, OsKCBP, OsCLASP, and OsMOR1 were cloned into the vector respectively, transformed into Agrobacterium tumefaciens strain, and expressed in Nicotiana benthamiana leaves. The results indicated that all of the MAPs were co-expressed with α-tubulin and localized to MTs, validating the availability and universality of the vector and that GL7, OsKCBP, OsCLASP, and OsMOR1 might be MAPs. The application of the co-expression vector constructed by us would facilitate studies on the interaction between tubulin/MT and MAP in tobacco transient expression systems or transgenic rice.

Assessing the impact of diverse mask types on COPD patients: a randomised controlled trial study protocol.

INTRODUCTION: Wearing masks has proven beneficial in preventing respiratory pathogen infections in individuals with chronic obstructive pulmonary disease (COPD). However, the impact of different mask types on physiological indicators and daily physical activity in COPD patients remains uncertain. This study aims to assess the immediate effects of various mask types on cardiopulmonary function indicators, subjective perceptions and the 6-minute walking distance (6MWD) in individuals with COPD. METHODS AND ANALYSIS: This randomised controlled trial will enrol 129 stable COPD patients. Participants will be randomly divided into three groups: control, N95 mask and surgical mask groups. Each group will undergo both a 6-minute seated test and a 6-minute walk test (6MWT), without or with their respective masks. A 10-minute interval will be provided between the two phases. The primary indicators of the study include the 6MWD and blood oxygen saturation. Secondary outcomes encompass blood pressure, pulse rate, Borg score, Rate of Perceived Exertion (RPE) score and subjective perception score. Oxygen saturation, pulse rate and blood pressure will be recorded four times during the trial, while Borg and RPE scores will be compared before and after the 6MWT. Additionally, subjective perception scores will be collected after each mask-wearing stage. ETHICS AND DISSEMINATION: This study has received approval from the Ethics Committee of the Affiliated Hospital of Gansu University of Chinese Medicine (approval number: 202335). We plan to disseminate research results through publication in a peer-reviewed journal or presentation at a conference. TRIAL REGISTRATION NUMBER: ChiCTR2300074554.

Electrochemical Properties of an Sn-Doped LATP Ceramic Electrolyte and Its Derived Sandwich-Structured Composite Solid Electrolyte.

An Li1.3Al0.3SnxTi1.7-x(PO4)3 (LATP-xSn) ceramic solid electrolyte was prepared by Sn doping via a solid phase method. The results showed that adding an Sn dopant with a larger ionic radius in a concentration of x = 0.35 enabled one to equivalently substitute Ti sites in the LATP crystal structure to the maximum extent. The uniform Sn doping could produce a stable LATP structure with small grain size and improved relative density. The lattice distortion induced by Sn doping also modified the transport channels of Li ions, which promoted the increase of ionic conductivity from 5.05 × 10-5 to 4.71 × 10-4 S/cm at room temperature. The SPE/LATP-0.35Sn/SPE composite solid electrolyte with a sandwich structure was prepared by coating, which had a high ionic conductivity of 5.9 × 10-5 S/cm at room temperature, a wide electrochemical window of 4.66 V vs. Li/Li+, and a good lithium-ion migration number of 0.38. The Li||Li symmetric battery test results revealed that the composite solid electrolyte could stably perform for 500 h at 60 °C under the current density of 0.2 mA/cm2, indicating its good interface stability with metallic lithium. Moreover, the analysis of the all-solid-state LiFePO4||SPE/LATP-0.35Sn/SPE||Li battery showed that the composite solid electrolyte had good cycling stability and rate performance. Under the conditions of 60 °C and 0.2 C, stable accumulation up to 200 cycles was achieved at a capacity retention ratio of 90.5% and a coulombic efficiency of about 100% after cycling test.

Nursing effect of Nasoscopically assisted nasogastri tube and nasojejunal tube placement.

OBJECTIVE: To investigate the nursing effect of nasoscopically assisted nasogastri tube and nasojejunal tube placement. METHODS: 94 patients who need to place nasogastric tube and nasojejunal tube to establish enteral nutrition were randomly divided into two groups: the observation group (n=49) and control group (n=45). The patients in the observation group received nasogastric tube placement and jejunal nutrition tube placement, and the patients in the control group received general gastroscope and placed gastric tube and jejunal nutrition tube through mouth. Success rate of catheterization, catheter pain score, satisfaction score, vital signs, completion time of catheterization, and complication were collected. RESULTS: the fluctuation of vital signs in control group was significantly higher than that in observation group. There was statistical significance between two groups in vital signs after intervention (P<0.05), mainly manifested in the heart rate, breathing and pulse pressure difference. On the other hand, there was no statistical significance between two groups in pulse oxygen after nursing intervention (P>0.05). The catheter pain score is obviously improved in the observation group compared with control group after intervention. The improvement score of satisfaction in the observation group was 91.47±7.65 points, and that in the control group was 83.64±5.24 points. The completion time of catheterization was improved in the observation group compared with control group. There was statistical significance between two groups in satisfaction score and completion time of catheterization (P<0.05). The rate of abdominal distention and diarrhea in the control group was higher than that in the observation group (P<0.05). CONCLUSION: Nasoscopically assisted nasogastri tube and nasojejunal tube placement has the advantages of simple and fast, short operation time, high success rate and few complications. It is the first choice of intubation method for enteral nutrition support treatment.

Effects of pulmonary rehabilitation training based on WeChat App on pulmonary function, adverse mood and quality of life of COVID-19 patients: A protocol for systematic review and meta-analysis.

BACKGROUND: Coronavirus disease 2019 (COVID-19) as a fatal epidemic has swept across the world, especially in India where the epidemic situation is the most serious. For COVID-19 patients, pulmonary rehabilitation training plays a significant role. However, it is still a controversial issue regarding the efficacy of WeChat APP-based pulmonary rehabilitation training in improving lung function, quality of life and bad mood of COVID-19 patients. To clarify this issue, a meta-analysis was conducted in this present study, so as to provide a basis for rehabilitation guidance of COVID-19 patients. METHODS: We systematically searched PubMed, medRxiv, Web of Science, Scopus, Chinese Science Citation Database, China National Knowledge Infrastructure, Chinese Biomedical Literature Database, Chinese Scientific Journal Database, and Wan-fang databases in May 2021 to identify randomized controlled trials and evaluate the effects of WeChat APP-based pulmonary rehabilitation training for COVID-19. Two researchers independently carried out data extraction. On the other hand, literature quality evaluation on the quality and meta-analysis of the included literature was performed with Revman5.3 software. RESULTS: The results of this meta-analysis will be submitted to a peer-reviewed journal for publication. CONCLUSION: This study will provide reliable evidence-based evidence on the effects of WeChat APP-based pulmonary rehabilitation training on lung function, bad mood, and quality of life in patients with COVID-19. ETHICS AND DISSEMINATION: Ethical approval was not required for this study. The systematic review will be published in a peer-reviewed journal, presented at conferences, and shared on social media platforms. OSF REGISTRATION NUMBER: DOI 10.17605/OSF.IO/MKXCH.

Anterior reduction and C1-ring osteosynthesis with Jefferson-fracture reduction plate (JeRP) via transoral approach for unstable atlas fractures.

BACKGROUND: To introduce a novel transoral instrumentation in the treatment of unstable fractures of the atlas. METHODS: From January 2008 to May 2018, 22 patients with unstable C1 fractures who received Jefferson-fracture reduction plate (JeRP) via transoral approach were retrospectively analyzed. The case history and the radiographs before and after surgery were noted. The type of fracture, the reduction of the fracture, and position of the internal fixation were assessed through preoperative and postoperative CT scans. RESULTS: All 22 patients successfully underwent anterior C1-ring osteosynthesis using the JeRP system, with a follow-up of 26.84 ± 9.23 months. Among them, 9 patients had transverse atlantal ligament (TAL) injury, including 3 in Dickman type I and 6 in type II. The preoperative lateral mass displacement (LMD) decreased from 7.13 ± 1.46 mm to 1.02 ± 0.65 mm after the operation. Bone union was achieved in all patients without implant failure or loss of reduction. There were no surgery-related complications, such as wound infection, neurological deficit, or vertebral artery injury. However, atlantoaxial dislocation occurred in 3 patients with Dickman type I TAL injury 3 months postoperatively without any neurological symptoms or neck pain. CONCLUSIONS: Transoral C1-ring osteosynthesis with JeRP is an effective surgical strategy to treat unstable atlas fractures with a safe, direct, and satisfactory reduction. The primary indication for the JeRP system is an unstable fracture (Gehweiler type I/III) or/ and TAL injury (Dickman type II).

Efficacy of a novel chelator BPCBG for removing uranium and protecting against uranium-induced renal cell damage in rats and HK-2 cells.

Chelation therapy is a known effective method to increase the excretion of U(VI) from the body. Until now, no any uranium chelator has been approved for emergency medical use worldwide. The present study aimed to evaluate the efficacy of new ligand BPCBG containing two catechol groups and two aminocarboxylic acid groups in decorporation of U(VI) and protection against acute U(VI) nephrotoxicity in rats, and further explored the detoxification mechanism of BPCBG for U(VI)-induced nephrotoxicity in HK-2 cells with comparison to DTPA-CaNa3. Chelating agents were administered at various times before or after injections of U(VI) in rats. The U(VI) levels in urine, kidneys and femurs were measured 24 h after U(VI) injections. Histopathological changes in the kidney and serum urea and creatinine and urine protein were examined. After treatment of U(VI)-exposed HK-2 cells with chelating agent, the intracellular U(VI) contents, formation of micronuclei, lactate dehydrogenase (LDH) activity and production of reactive oxygen species (ROS) were assessed. It was found that prompt, advanced or delayed injections of BPCBG effectively increased 24 h-urinary U(VI) excretion and decreased the levels of U(VI) in kidney and bone. Meanwhile, BPCBG injection obviously reduced the severity of the U(VI)-induced histological alterations in the kidney, which was in parallel with the amelioration noted in serum indicators, urea and creatinine, and urine protein of U(VI) nephrotoxicity. In U(VI)-exposed HK-2 cells, immediate and delayed treatment with BPCBG significantly decreased the formation of micronuclei and LDH release by inhibiting the cellular U(VI) intake, promoting the intracellular U(VI) release and inhibiting the production of intracellular ROS. Our data suggest that BPCBG is a novel bi-functional U(VI) decorporation agent with a better efficacy than DTPA-CaNa3.

Efficacy of Chelator CBMIDA-CaNa2 for the Removal of Uranium and Protection against Uranium-induced Cell Damage in Human Renal Proximal Tubular Cells.

ABSTRACT: In animal experiments, catechol-3,6-bis(methyleiminodiacetic acid) (CBMIDA) was proven to be an effective chelator for the decorporation of uranium (U)(VI). In the present study, the authors investigated the molecular processes of CBMIDA-CaNa2 on the removal of U(VI) at the cellular level and explored its protective effects and mechanism against U(VI)-induced cell damage in HK-2 human renal proximal tubular cells. The results indicated that the chelating U(VI) effect of CBMIDA-CaNa2 was superior compared to that of DTPA-CaNa3; more specifically, at concentrations of 50 and 250 µM, CBMIDA-CaNa2 can significantly reduce U(VI) uptake and increase U(VI) release in U(VI)-exposed HK-2 cells after immediate or 24-h and 48-h delayed chelator administration better than those of DTPA-CaNa3. Furthermore, CBMIDA-CaNa2 significantly decreased the lactate dehydrogenase release and the formation of micronuclei and inhibited the production of intracellular reactive oxygen species (ROS) in HK-2 cells exposed to U(VI), whereas DTPA-CaNa3 was demonstrated to be ineffective. By reviewing the results of animal experiments conducted by several other investigators, including this lab, the authors found that removal efficacy and protective effects of these two chelators for U(VI) at the cellular level agreed well with those of animal studies. In addition, although U(VI) induced the increase of metallothionein protein expression in HK-2 cells, CBMIDA-CaNa2 can mobilize and remove the U(VI) from metallothionen (MT) after 48-h delayed chelator treatment. These results suggested that CBMIDA-CaNa2 protected against U(VI)-induced HK-2 cells damaged by reducing U(VI) uptake, increasing U(VI) release and scavenging the U(VI)-induced intracellular ROS.

Combination effects of chronic cadmium exposure and gamma-irradiation on the genotoxicity and cytotoxicity of peripheral blood lymphocytes and bone marrow cells in rats.

This work investigated the effects of chronic cadmium (Cd) exposure combined with γ-ray irradiation on the cytotoxicity and genotoxicity of peripheral blood cells and bone marrow cells in rats. Results showed that when the rats were exposed to low dose (LD) Cd of 0.1mg CdCl2/(kgd) for 8 and 12 weeks, the Cd concentration in blood reached to 135-140 µg/L and no toxic effects on peripheral blood lymphocytes, white blood cells (WBC) and granulocyte-monocyte (GM) progenitor cells were observed except polychromatic erythrocytes (PCE) of bone marrow. Moreover, this chronic LD Cd exposure significantly decreased irradiation-induced micronucleus (MN) formation and hypoxanthine-guanine phosphoribosyl transferase (hprt) mutation in lymphocytes and PCE, while the combination of LD Cd exposure and irradiation induced the additive metallothionein (MT) protein expression in bone marrow cells. When the rats were exposed to a high dose (HD) Cd of 0.5mg CdCl/2(kgd) for 8 and 12 weeks, the blood Cd level approached to 458-613 µg/L and an inflammatory response was induced, meanwhile, MN formation and hprt mutation were markedly increased, and the ratio of PCE/NCE (normochromatic erythrocyte) was significantly decreased. Furthermore, when the rats were exposed to HD Cd plus 2 Gy irradiation, additive toxic effects on MN formation, hprt mutation, PCE damage and GM progenitor cell proliferation were observed, while this combination treatment resulted in an obvious reduction of MT protein compared to HD Cd group. In conclusion, chronic exposure to LD Cd induced the adaptive response to irradiation in the genotoxicity of peripheral blood lymphocytes and PCE of bone marrow by the up-regulation of Cd-induced MT protein, but the combination of HD Cd exposure and irradiation generated the additive effects on the cytotoxicity and genotoxicity in peripheral blood lymphocytes and bone marrow cells.

[Effect of the chelator BPCBG on the decorporation of uranium in vivo and uranium-induced damage of human renal tubular epithelial cells in vitro].

This study is to assess the efficacy of BPCBG on the decorporation of uranium (VI) and protecting human renal proximal tubular epithelial cells (HK-2) against uranium-induced damage. BPCBG at different doses was injected intramuscularly to male SD rats immediately after a single intraperitoneal injection of UO2(CH3COO)2. Twenty-four hours later uranium contents in urine, kidneys and femurs were measured by ICP-MS. After HK-2 cells were exposed to UO2(CH3COO)2 immediately or for 24 h followed by BPCBG treatment at different doses for another 24 or 48 h, the uranium contents in HK-2 cells were measured by ICP-MS, the cell survival was assayed by cell counting kit-8 assay, formation of micronuclei was determined by the cytokinesis-block (CB) micronucleus assay and the production of intracellular reactive oxygen species (ROS) was detected by 2',7'-dichlorofluorescin diacetate (DCFH-DA) oxidation. DTPA-CaNa3 was used as control. It was found that BPCBG at dosages of 60, 120, and 600 micromol kg(-1) resulted in 37%-61% increase in 24 h-urinary uranium excretion, and significantly decreased the amount of uranium retention in kidney and bone to 41%-31% and 86%-42% of uranium-treated group, respectively. After HK-2 cells that had been pre-treated with UO2(CH3COO)2 for 24 h were treated with the chelators for another 24 h, 55%-60% of the intracellular uranium was removed by 10-250 micromol L(-1) of BPCBG. Treatment of uranium-treated HK-2 cells with BPCBG significantly enhanced the cell survival, decreased the formation of micronuclei and inhibited the production of intracellular ROS. Although DTPA-CaNa3 markedly reduced the uranium retention in kidney of rats and HK-2 cells, its efficacy of uranium removal from body was significantly lower than that of BPCBG and it could not protect uranium-induced cell damage. It can be concluded that BPCBG effectively decorporated the uranium from UO2(CH3COO)2-treated rats and HK-2 cells, which was better than DTPA-CaNa3. It could also scavenge the uranium-induced intracellular ROS and protect against the uranium-induced cell damage. BPCBG is worth further investigation.

The role of nucleophosmin/B23 in radiation-induced chromosomal instability in human lymphoblastoid cells of different p53 genotypes.

PURPOSE: To investigate the role of nucleophosmin (NPM/B23) in radiation-induced chromosomal instability and apoptosis in human lymphoblastoid cells with different protein 53 (p53) status. MATERIALS AND METHODS: Wild type (wt) p53 TK6 and mutant type (mt) p53 WTK1 with or without short hairpin RNA (shRNA)-mediated silencing of NPM, TK6 with or without short interfering RNA (siRNA)-mediated silencing of p53 (p53i and NEGi) were irradiated with 4 Gy gamma-rays. Six to 48 h after irradiation, the index of apoptosis, chromosome aberration, cell cycle distribution and the levels of total NPM and phosphorylated-threonine 199 (pThr¹99) NPM proteins were measured. Cells in some dishes were treated with 10 µM Olomoucine (OLO) for 3 h before irradiation and remained in the medium after irradiation. RESULTS: The rates of radiation-induced apoptosis in TK6 and TK6/NEGi were about 2-fold of those in WTK1 and TK6/p53i, while the frequencies of polyploidy in TK6 and TK6/NEGi were obviously lower than those in WTK1 and TK6/p53i. Moreover, after irradiation, pThr¹99 NPM levels increased significantly in WTK1 and TK6/p53i, and slightly increased in TK6 and TK6/NEGi, indicating that the increased level of pThr¹99 NPM was related to p53 status. When Thr¹99 hyperphosphorylation of NPM was inhibited by OLO or when NPM was knocked down, we found that radiation-induced apoptosis was more pronounced and polyploidy formation was reduced as compared with negative control while the magnitude of these changes in TK6 was obviously higher than that in WTK1, indicating that NPM has an antagonistic interaction with wt p53. CONCLUSIONS: NPM/B23 plays an important role in protecting cells from radiation-induced apoptosis and increasing polyploidy formation via either a p53 or non-p53 pathway.