A Statistical Evaluation of Uncoupling Protein 1 in the Limited Area of Brown Adipose Tissue by Immunoelectron Microscopy

Uncoupling protein 1 (UCP1) expressed by the brown adipose tissue (BAT) in the mitochondrial crista acts as a homeostatic thermogenerator of eutherians. The evaluation of UCP1 expression in the BAT offers significant scientific insight, especially in studies targeting limited areas such as the periarterial and pericardial regions of small experimental mammals. However, the negligible amount of this adipose tissue would render the general quantitative evaluation of the protein unreliable because of lipid contamination and low protein concentration. To address this problem, we quantitatively evaluated UCP1 expression in the mitochondrion of the mouse interscapular BAT using immunoelectron microscopy and immunohistochemical studies using a combination of primary and secondary antibodies in scheme A (rabbit anti-UCP1 IgG/gold particle-conjugated goat anti-rabbit IgG), B (rabbit IgG/gold particle-conjugated goat anti-rabbit IgG), C (rabbit anti-UCP1 IgG/gold particle-unconjugated goat anti-rabbit IgG), and D (rabbit IgG/gold particle-unconjugated goat anti-rabbit IgG). Scheme A shows the immunopositive reaction of obvious gold particles in the mitochondrial area, whereas other procedures revealed less distinctive reactions. The distinctive gold particle immunoreaction comprised electrical high-density spots with a mean diameter of >5 nm. However, in scheme B, the electrical high-density spots were scattered outside the mitochondrion and were significantly smaller than 4 nm;schemes C and D demonstrated few immunoreactions. Logistic regression analysis between schemes A and B showed that the threshold diameter of the electrical high-density spots measuring >5 nm indicated a true positive immunoreaction to anti-UCP1 antibody specifically in the mitochondrial area. Minor statistical difference was observed in the primary anti-UCP1 antibody between polyclonal IgG and monoclonal antibodies. Therefore, immunoelectron microscopy might be useful for evaluating negligible protein expression in some limited areas, such as UCP1 expression in the BAT of small experimental animals.

Food synthetic biology-driven protein supply transition: From animal-derived production to microbial fermentation

Animal-derived protein production is one of the major traditional protein supply methods,which continues to face increasing challenges to satisfy global needs due to population growth,augmented individual protein consumption,and aggravated environmental pollution.Thus,ensuring a sustainable protein source is a considerable challenge.The emergence and development of food synthetic biology has enabled the establishment of cell factories that effectively synthesize proteins,which is an important way to solve the protein supply problem.This review aims to discuss the existing problems of traditional protein supply and to elucidate the feasibility of synthetic biology in the process of protein synthesis.Moreover,using artificial bioengineered milk and artificial bioengineered eggs as examples,the progress of food protein supply transition based on synthetic biology has been systematically summarized.Additionally,the future of food synthetic biology as a potential source of protein has been also discussed.By strengthening and innovating the application of food synthetic biology technologies,including genetic engineering and high-throughput screening methods,the current limitations of artificial foods for protein synthesis and production should be addressed.Therefore,the development and industrial production of new food resources should be explored to ensure safe,high-quality,and sustainable global protein supply.

Preparation of γ-Al2O3 via Hydrothermal Synthesis Using ρ-Al2O3 as Raw Material for Propane Dehydrogenation

γ-Al2O3 was prepared by hydrothermal synthesis usingρ-Al2O3 and urea as raw materials.In this work,the eff ects of the molar ratio of CO(NH2)2/Al and reaction temperature were investigated,and a Pt–Sn–K/γ-Al2O3 catalyst was prepared.The ammonium aluminum carbonate hydroxide(AACH),γ-Al2O3,and Pt–Sn–K/γ-Al2O3 were characterized by X-ray diff raction,scanning electron microscopy,transmission electron microscopy,N2 adsorption–desorption,thermogravimetry–differential thermal analysis,and NH3 temperature-programmed desorption techniques.The reactivity of Pt–Sn–K/γ-Al2O3 for propane dehydrogenation was tested in a micro-fixed-bed reactor.The results show thatγ-Al2O3 with a specific surface area of 358.1 m 2/g and pore volume of 0.96 cm 3/g was obtained when the molar ratio of CO(NH2)2/Al was 3:1 and the reaction temperature was 140℃.The alumina obtained by calcination of AACH has a higher specific surface area and larger pore volume than the industrial pseudo-boehmite does.The catalyst prepared from AACH as precursor showed high selectivity and conversion,which can reach 96.1%and 37.6%,respectively,for propane dehydrogenation.

Paenarthrobacter nitroguajacolicus can promote cucumber growth and control cucumber corynespora leaf spot

Currently,the disease control in cucumber mainly depends on agrochemicals,which is not an environmentally benign strategy.Biocontrol bacteria not only resist plant pathogens but also promote plant growth,which is ecofriendly and sustainable option.A biocontrol bacterial strain BJ-5 was screened using Corynespora cassiicola as the target pathogen,and BJ-5 was determined to be Paenarthrobacter nitroguajacolicus by morphological and molecular methods.The effect of BJ-5 on C.cassiicola was studied,including the spore germination,cell membrane permeability and infected cucumbers.BJ-5 inhibited the germination of C.cassiicola spores in vitro and led to atrophy and deformation of the C.cassiicola budding tubes.BJ-5 caused the relative extracellular conductivity of C.cassiicola mycelia to increase compared with the control.Additionally,BJ-5 reduced the severity of cucumber corynespora leaf spot of cucumber infected with C.cassiicola.The inhibition efficacy of BJ-5 suspension as a foliar spray against cucumber corynespora leaf spot reached 63%inhibition,which is higher than a 5000-fold dilution of Luna-Son SC fungicide.In addition,BJ-5 was tested on the emergence of cucumber seedlings,recording the biomass and photosynthesis of cucumber during the growth period.BJ-5 at 1.5×10^(5)CFU·mL^(−1)promoted the germination of cucumber seeds and increased biomass and photosynthesis at the adult plant stage.Also,the secondary metabolites of BJ-5 were determined.BJ-5 could produce chitinases,siderophore,cellulase,amylase and protease in the respective medium.Finally,adaptation assay of BJ-5 showed good salt tolerance and good adaptability in alkaline conditions,and that BJ-5 retains inhibition of fungi activity at higher temperatures.This is the first report of the biocontrol by P.nitroguajacolicus with antagonism to C.cassiicola and promote cucumber growth.This study indicates that P.nitroguajacolicus may serve as potential biocontrol agents against cucumber corynespora leaf spot fungus.

Metabolic engineering of Escherichia coli for the production of Lacto-N-neotetraose(LNnT)

Lacto-N-neotetraose(LNnT),one of the most important human milk oligosaccharides,can be used as infants’food addi-tives.Nowadays,extraction,chemical and biological synthesis were utilized to obtain LNnT,while these methods still face some problems such as low yield and high cost.The aim of current work is to construct a de novo biosynthesis pathway of LNnT in E.coli K12 MG1655.The lgtA and lgtB were first expressed by a plasmid,resulting in a LNnT titer of 0.04 g/L.To improve the yield of LNnT on substrate lactose,lacZ and lacI were knocked out,and lacY was over-expressed.As a result,the yield of LNnT on lactose increased from 0.01 to 0.09 mol/mol,and the titer of LNnT elevated to 0.41 g/L.In addition,the pathway was regulated using the titer of Lacto-N-triose II(LNTII)as a measure,and obtained a high titer strain of LNnT for 1.04 g/L.Finally,the gene expressions were fine-tuned,the titer of LNnT reached 1.2 g/L,which was 93%higher than the control strain,and the yield on lactose reached 0.28 mol/mol.The engineering strategy of pathway construction and modulation used in this study is applicable to facilitate the microbial production of other metabolites in E.coli.

DNA methylation changes induced by BDE-209 are related to DNA damage response and germ cell development in GC-2spd

Decabrominated diphenyl ether(BDE-209)is generally utilized in multiple polymer materials as common brominated flame retardant.BDE-209 has been listed as persistent organic pollutants(POPs),which was considered to be reproductive toxin in the environment.But it still remains unclear about the effects of BDE-209 on DNA methylation and the inducedmale reproductive toxicity.Due to the extensive epigenetic regulation in germ line development,we hypothesize that BDE-209 exposure impacts the statue of DNA methylation in spermatocytes in vitro.Therefore,the mouse GC-2spd(GC-2)cells were used for the genome wide DNA methylation analysis after treated with 32μg/mL BDE-209 for 24 hr.The results showed that BDE-209 caused genomic methylation changes with 32,083 differentially methylated CpGs in GC-2 cells,including 16,164(50.38%)hypermethylated and 15,919(49.62%)hypomethylated sites.With integrated analysis ofDNAmethylation data and functional enrichment,we found that BDE-209 might affect the functional transcription in cell growth and sperm development by differential gene methylation.qRT-PCR validation demonstrated the involvement of p53-dependent DNA damage response in the GC-2 cells after BDE-209 exposure.In general,our findings indicated that BDE-209-induced genome wide methylation changes could be interrelated with reproductive dysfunction.This study might provide new insights into the mechanisms of male reproductive toxicity under the environmental exposure to BDE-209.