[Effects of Methionine Restriction on Proliferation, Cell Cycle, and Apoptosis of Human Acute Leukemia Cells].

OBJECTIVE: To investigate the effects of methionine restriction on proliferation, cell cycle and apoptosis of human acute leukemia cells. METHODS: Cell Counting Kit-8 (CCK-8) assay was used to detect the effect of methionine restriction on HL-60 and Jurkat cells proliferation. The effect of methionine restriction on cell cycle of HL-60 and Jurkat cells was examined by PI staining. Annexin V-FITC / PI double staining was applied to detect apoptosis of HL-60 and Jurkat cells following methionine restriction. The expression of cell cycle-related proteins cyclin B1, CDC2 and apoptosis-related protein Bcl-2 was evaluated by Western blot assay. RESULTS: Methionine restriction significantly inhibited the proliferation of HL-60 and Jurkat cells in a time-dependent manner (HL-60: r =0.7773, Jurkat: r =0.8725), arrested the cells at G2/M phase (P < 0.001), and significantly induced apoptosis of HL-60 and Jurkat cells (HL-60: P < 0.001; Jurkat: P < 0.05). Furthermore, Western blot analysis demonstrated that methionine restriction significantly reduced the proteins expression of Cyclin B1 (P < 0.05), CDC2 (P < 0.01) and Bcl-2 (P < 0.001) in HL-60 and Jurkat cells. CONCLUSION: Acute leukemia cells HL-60 and Jurkat exhibit methionine dependence. Methionine restriction can significantly inhibit the proliferation, promote cell cycle arrest and induce apoptosis of HL-60 and Jurkat cells, which suggests that methionine restriction may be a potential therapeutic strategy for acute leukemia.

Aurantio-obtusin ameliorates obesity by activating PPARα-dependent mitochondrial thermogenesis in brown adipose tissues.

Obesity contributes to the progression of various chronic diseases, and shortens life expectancy. With abundant mitochondria, brown adipose tissue (BAT) dissipates energy through heat to limit weight gain and metabolic dysfunction in obesity. Our previous studies have shown that aurantio-obtusin (AO), a bioactive ingredient in Chinese traditional medicine Cassiae semen significantly improves hepatic lipid metabolism in a steatotic mouse model. In the current study we investigated the effects of AO on lipid metabolism in the BAT of diet-induced obesity mice and in oleic acid and palmitic acid (OAPA)-stimulated primary mature BAT adipocytes. Obese mice were established by feeding a HFHS diet for 4 weeks, and then administered AO (10 mg/kg, i.g.) for another 4 weeks. We showed that AO administration significantly increased the weight of BAT and accelerated energy expenditure to protect the weight increase in the obese mice. Using RNA sequencing and molecular biology analysis we found that AO significantly enhanced mitochondrial metabolism and UCP1 expression by activating PPARα both in vivo and in vitro in the primary BAT adipocytes. Interestingly, AO administration did not improve metabolic dysfunction in the liver and white adipose tissue of obese mice after interscapular BAT excision. We demonstrated that low temperature, a trigger of BAT thermogenesis, was not a decisive factor for AO to stimulate the growth and activation of BATs. This study uncovers a regulatory network of AO in activating BAT-dependent lipid consumption and brings up a new avenue for the pharmaceutical intervention in obesity and related comorbidities.

[Effect of a novel phosphodiesterase 5 inhibitor, CPD1, on renal interstitial fibrosis after unilateral renal ischemia-reperfusion injury].

This study was designed to evaluate the protective effect of CPD1, a novel phosphodiesterase 5 inhibitor, on renal interstitial fibrosis after unilateral renal ischemia-reperfusion injury (UIRI). Male BALB/c mice were subjected to UIRI, and treated with CPD1 once daily (i.g, 5 mg/kg). Contralateral nephrectomy was performed on day 10 after UIRI, and the UIRI kidneys were harvested on day 11. Hematoxylin-eosin (HE), Masson trichrome and Sirius Red staining methods were used to observe the renal tissue structural lesions and fibrosis. Immunohistochemical staining and Western blot were used to detect the expression of proteins related to fibrosis. HE, Sirius Red and Masson trichrome staining showed that CPD1-treated UIRI mice had lower extent of tubular epithelial cell injury and deposition of extracellular matrix (ECM) in renal interstitium compared with those in the fibrotic mouse kidneys. The results from immunohistochemistry and Western blot assay indicated significantly decreased protein expressions of type I collagen, fibronectin, plasminogen activator inhibitor-1 (PAI-1) and α-smooth muscle actin (α-SMA) after CPD1 treatment. In addition, CPD1 dose-dependently inhibited the expression of ECM-related proteins induced by transforming growth factor ß1 (TGF-ß1) in normal rat kidney interstitial fibroblasts (NRK-49F) and human renal tubular epithelial cell line (HK-2). In summary, the novel PDE inhibitor, CPD1, displays strong protective effects against UIRI and fibrosis by suppressing TGF-ß signaling pathway and regulating the balance between ECM synthesis and degradation through PAI-1.

Emission of volatile organic compounds (VOCs) from application of commercial pesticides in China.

Applying pesticides can result in emissions of volatile organic compounds (VOCs), but little is known about VOC emission characteristics and the quantities in particular regions. We investigated the use of pesticides in China based on a large-scale survey of 330 counties in 31 provinces and evaluated the national pesticide VOC emission potentials based on thermogravimetric analysis of 1930 commercial pesticides. The results showed that herbicides were the most extensively used pesticide category in China, accounting for 43.47%; emulsifiable concentrate (EC), suspension concentrate, and wettable powder were the dominant pesticide formulations, with proportions of 26.75%, 17.68%, and 17.31%, respectively. The VOC emission potential coefficient (EP) of the liquid formulations was higher than the solid formulations, and the maximum mean EP was 45.59% for EC and the minimum was 0.76% for WP. Among 437 high-VOC pesticide products used in China, EC accounted for 83.52%, and 16.93% of those contained abamectin. The total VOC emissions derived from commercial pesticides in China were 280 kt (kilotons) in 2018, and 65.35% of the contribution was derived from EC. Shandong, Hunan, and Henan were the three provinces with the highest pesticide VOC emissions (>21 kt/y). The emission rate of VOCs from pesticides was 24.80 t/d in China, which was higher than in San Joaquin Valley, California. These findings suggest that some comprehensive measures (e.g., perfecting pesticide management policy, strict supervision for pesticide production and use, and strengthening pesticide reduction publicity) should be taken to reduce VOC emissions from pesticide applications.

[The Effect of Novel Phosphodiesterase 4 Inhibitor ZL-n-91 to the Proliferation of Leukemia Cells].

OBJECTIVE: To investigate the inhibitory effects of novel phosphodiesterase 4 inhibitor ZL-n-91 to the proliferation of leukemia cells L1210 and K562. METHODS: CCK-8 method was used to detect the effect of ZL-n-91 to the proliferation of L1210 and K562 cells, and the proliferation rate, IC50 were calculated. The effects of ZL-n-91 to the cycle of L1210 and K562 cells was detected by PE single staining, and the effects of ZL-n-91 to the apoptosis of L1210 and K562 cells was detected by PE/7AA-D double staining. Western blot was used to detect the effect of ZL-n-91 to the expression levels of apoptosis related proteins. Subcutaneous tumor transplantation model of acute lymphoblastic leukemia L1210 was established in the nude mice, and the inhibitory effect of oral administration of ZL-n-91 to the xenograft was observed. RESULTS: ZL-n-91 showed a significant inhibitory effect to the proliferation of leukemia cells L1210 and K562 in a dose-dependent manner (P<0.001). After treated by ZL-n-91, the leukemia cells L1210 and K562 in the S-phase in cell cycle decreased significantly compared with those in control group (P<0.01). The apoptosis of leukemia cells L1210 and K562 could be induced by ZL-n-91 (P<0.001), and the expression level of apoptosis related protein BAX significantly increased. In the animal experiment, the result showed that ZL-n-91 could significantly inhibit the growth of subcutaneously transplantation tumor (P<0.05). CONCLUSION: The novel phosphodiesterase 4 inhibitor ZL-n-91 can effectively inhibit the proliferation of leukemia cells L1210 and K562, which has the potential of anti-leukemia drug development.

[Effects of Water Supply Quality and Social Activity on the Microbial Community in Baiyangdian Wetland].

To clarify the impact of water quality and social activity in the Baiyangdian wetland on the biological community, the change characteristics of bacterial, fungal, and archaeal communities in different areas of the Dian District were studied. Samples were collected at the entrance of Fuhe District (NBB), tourist areas with frequent human social activities (NBD), residential breeding areas (NBX), and village sparse areas (NBN). The physical and chemical characteristics and biological communities of the samples were evaluated. The results of the study show that the COD concentration of organic pollutants in the NBB was 12.35 mg·L-1, and the total nitrogen concentration was 10.12 mg·L-1, that the concentration was highest. Moreover, the water quality in NBD and NBX was better than that of NBB. The NBN area exhibited the best water quality, with COD and total nitrogen concentration values of 6.9 mg·L-1 and 1.82 mg·L-1, respectively. Many types of NBB bacteria were recorded, with a diversity index of 5.86, and NBN diversity index exceeding 4.78. The dominant bacterial flora in all samples was the Proteobacteria, which accounts for 68.8% of the total bacterial communities in NBN samples. The diversity index of fungi in NBB was only 2.14. There were many types of fungi in NBN, with a diversity index of 3.23. Chytridiomycota was found in the NBD and NBN, accounting for 5.4% and 9.8% of the total number of fungi, respectively. The Chytridiomycota was main decomposer of hard to degrade organic carbon. The diversity of archaea of NBN was the lowest among all the samples. Crenarchaeota was the dominant phylum, which accounts for 39.0%, 51.9%, 47.3%, and 30.1% of NBB, NBD, NBX, and NBN samples, respectively. The number of Halobacterota was lower than Crenarchaeota. The main factor of eutrophication and microbial community changes in Baiyangdian wetland was the results of the combined action of external and internal pollution. Both external and internal pollution increased the organic matter, nitrogen and phosphorus content in the water, and the microbial community structure has changed significantly. The contents of organic matter, nitrogen, and phosphorus in water were increased and the microbial community structures were changed significantly by the increase of both external and internal pollution.

[Purification Characteristics of Urban Tail Water from Sewage Treatment Plant by Biofilm Ecological Floating Bed].

The purification characteristics of nitrogenous tail water were investigated using a biofilm ecological floating bed technology to study biofilm length, hydraulic retention time (HRT), and biofilm coverage area with the aim to explore the purification characteristics of biofilm on urban tail water. Results show that the removal rates of NH4+-N, NO3--N, and TN were 90.82%, 62.7%, and 81.96%, respectively, at the half water depth to the suspended biofilm. The removal rate of NH4+-N was only 22.07%, and the concentration changes of NO3--N and TN were not obvious throughout the whole water depth to the suspended biofilm. When the HRT was 6 days, the removal rates of NH4+-N and TN could reach 82.01% and 62.88%, respectively, whereas the lowest rates were 55.24% and 46.82%, respectively. When the HRT was 12 days, the removal rates of NH4+-N and TN reached up to 81.4% and 79.93%, respectively, whereas the lowest rates were 8.73% and 17.23%, respectively. In contrast, the nitrogen removal efficiency was high and stable when the HRT was 6 days. When the coverage area was 10%, the removal rate was decreased in one operation cycle. When the coverage area was 20%, the removal rate showed an upward trend. Under the conditions of 10% and 20% biofilm coverage area, the removal rates of TN were 62.88% and 71.09%, respectively.

Impairment of spermatogenesis and sperm motility by the high-fat diet-induced dysbiosis of gut microbes.

OBJECTIVE: High-fat diet (HFD)-induced metabolic disorders can lead to impaired sperm production. We aim to investigate if HFD-induced gut microbiota dysbiosis can functionally influence spermatogenesis and sperm motility. DESIGN: Faecal microbes derived from the HFD-fed or normal diet (ND)-fed male mice were transplanted to the mice maintained on ND. The gut microbes, sperm count and motility were analysed. Human faecal/semen/blood samples were collected to assess microbiota, sperm quality and endotoxin. RESULTS: Transplantation of the HFD gut microbes into the ND-maintained (HFD-FMT) mice resulted in a significant decrease in spermatogenesis and sperm motility, whereas similar transplantation with the microbes from the ND-fed mice failed to do so. Analysis of the microbiota showed a profound increase in genus Bacteroides and Prevotella, both of which likely contributed to the metabolic endotoxaemia in the HFD-FMT mice. Interestingly, the gut microbes from clinical subjects revealed a strong negative correlation between the abundance of Bacteroides-Prevotella and sperm motility, and a positive correlation between blood endotoxin and Bacteroides abundance. Transplantation with HFD microbes also led to intestinal infiltration of T cells and macrophages as well as a significant increase of pro-inflammatory cytokines in the epididymis, suggesting that epididymal inflammation have likely contributed to the impairment of sperm motility. RNA-sequencing revealed significant reduction in the expression of those genes involved in gamete meiosis and testicular mitochondrial functions in the HFD-FMT mice. CONCLUSION: We revealed an intimate linkage between HFD-induced microbiota dysbiosis and defect in spermatogenesis with elevated endotoxin, dysregulation of testicular gene expression and localised epididymal inflammation as the potential causes. TRIAL REGISTRATION NUMBER: NCT03634644.

[Characteristics of Nitrogen Metabolism by Denitrifying Bacterial Agents in Low Dissolved Oxygen Black Odor Water].

In order to deal with low dissolved oxygen black odor water body of Guangdong-Hong Kong-Macao Greater Bay Area, the nitrogen metabolic capacity of ammoniated bacteria, nitrifying bacteria and denitrifying bacteria under low dissolved oxygen concentration was studied, and the relationship between denitrification efficiency and nitrogen conversion of different bacteria was clarified, which provided theoretical basis for the treatment of black odor water of Guangdong-Hong Kong-Macao Greater Bay Area. Nitrogen conversion characteristics of ammoniated bacteria (Staphylococcus sp. Ay), nitrifying bacteria (Microbacterium sp. Xw), and denitrifying bacteria (Arthrobacter sp. Fy) in simulated wastewater were detected by different concentration of bacteria and different concentration of substrates. The nitrogen metabolism of Ay, Xw and Fy in black odor water at the same concentration under low dissolved oxygen condition and the characteristics of nitrogen metabolism in different concentrations of black odor water were studied. The results showed that the bacterial concentration of Ay bacterium agent increased three times, but the ammoniation efficiency did not multiply. It indicated that increasing the concentration of ammoniated bacterium was not the best way to increase the ammoniation rate. Xw bacteria have a significant effect on the change of nitrate concentration. At 60 h, the nitrate concentration was increased 180% at low bacterial concentration, was 231% at a high bacterial concentration. Fy denitrification efficiency was high, that nitrate nitrogen can be removed quickly at lower concentration. Appropriate increase of total nitrogen concentration can increase the ammoniation efficiency of Ay microbial agent, but too high would inhibit the nitrogen metabolism efficiency of Ay microbial agent. The concentration of nitrate nitrogen of Xw bacterium reagent was increased (3736%) when the total nitrogen was 25 mg·L-1. The Fy microbial agent has no obvious effect on ammonia nitrogen removal. When the total nitrogen concentration was 25 mg·L-1, the removal rate of nitrate by Fy bacterium was the best. Overall, the results showed that the concentration of ammoniated bacteria and denitrifying bacteria could meet the need of nitrogen metabolism, and the nitrifying bacteria could increase the nitrate nitrogen synthesis rate with the increase of the concentration of nitrifying bacteria. In a certain range, increasing the concentration of substrate could increase the efficiency of nitrogen metabolism of bacterial agents, and excessive concentration would inhibit the effect of nitrogen metabolism. The study provides data support for the problem of black odor water in Guangdong-Hong Kong-Macao Greater Bay Area.

Genetic variations of human CYP2D6 in the Chinese Han population.

AIM: The purpose of this study was to determine the genetic polymorphisms of the CYP2D6 gene and to elucidate the allele distribution pattern in the Chinese Han population. MATERIALS & METHODS: We used PCR and bidirectional sequencing methods to analyze all nine exons of the CYP2D6 gene in 2129 unrelated, healthy Chinese Han subjects from two geographical locations in China: the northern and southern regions. RESULTS: In total, 165 mutated sites were detected in 2129 participants, of which 67 sites were reported for the first time. Among these novel mutation sites, 22 were nonsynonymous and 12 were named as novel alleles (*87-*93, *94A, *94B and *95-*98) by the Human CYP Allele Nomenclature Committee. In addition, 29 previously reported alleles and 84 genotypes were also detected in 1954 volunteers. Functional prediction of novel variants revealed that eight variants might have a deleterious effect on CYP2D6. Linkage disequilibrium analysis and tagSNP selection were performed separately. By using these methods, distinct differences were found between the two regions. CONCLUSION: This study provides the most comprehensive data concerning CYP2D6 polymorphisms in the Chinese Han population to date and increases the number of known alleles; these findings may greatly contribute to the development of personalized medicine for the Chinese Han population.