Effects of alcohol on the composition and metabolism of the intestinal microbiota among people with HIV: a cross-sectional study.

OBJECTIVES: Alcohol consumption is not uncommon among people with HIV (PWH) and may exacerbate HIV-induced intestinal damage, and further lead to dysbiosis and increased intestinal permeability. This study aimed to determine the changes in the faecal microbiota and its association with alcohol consumption in HIV-infected patients. METHODS: A cross-sectional survey was conducted between November 2021 and May 2022, and 93 participants were recruited. To investigate the alterations of alcohol misuse on fecal microbiology in HIV-infected individuals, we performed 16s rDNA gene sequencing on fecal samples from the low to moderate drinking (n=21) and non-drinking (n=72) groups. RESULTS: Comparison between groups using alpha and beta diversity showed that the diversity of stool microbiota in the low to moderate drinkinge group did not differ from that of the non-drinking group (all P>0.05). The Linear discriminant Analysis effect size (LEfSe) algorithm was to determine the bacterial taxa associated with alcohol consumption, and the results showed altered fecal bacterial composition in HIV-infected patients who consumed alcohol, with Coprobacillus, Pseudobutyrivibrio and Peptostreptococcaceae enriched, and Pasteurellaceae and Xanthomonadaceae were depleted. In addition, by using the Kyoto Encyclopedia of Genes and Genomes (KEGG) functional microbiome features were also found to be altered in the low to moderate drinking group, showing a reduction in metabolic pathways (P=0.036) and cardiovascular disease pathway (P=0.006). CONCLUSION: Low to moderate drinking will change the composition, metabolism and cardiovascular disease pathway of the gut microbiota of HIV-infected patients.

Metabolic Signature of Healthy Lifestyle and Risk of Rheumatoid Arthritis: Observational and Mendelian Randomization Study.

BACKGROUND: Although substantial evidence reveals that healthy lifestyle behaviors are associated with a lower risk of rheumatoid arthritis (RA), the underlying metabolic mechanisms remain unclear. OBJECTIVES: This study aimed to identify the metabolic signature reflecting a healthy lifestyle and investigate its observational and genetic linkage with RA risk. METHODS: This study included 87,258 UK Biobank participants (557 cases with incident RA) aged 37-73 y with complete lifestyle, genotyping, and nuclear magnetic resonance (NMR) metabolomics data. A healthy lifestyle was assessed based on 5 factors: healthy diet, regular exercise, not smoking, moderate alcohol consumption, and normal body mass index. The metabolic signature was developed by summing the selected metabolites' concentrations weighted by the coefficients using elastic net regression. We used the multivariate Cox model to assess the associations between metabolic signatures and RA risk, and examined the mediating role of the metabolic signature in the impact of a healthy lifestyle on RA. We performed genome-wide association analysis (GWAS) to obtain genetic variants associated with the metabolic signature and then conducted Mendelian randomization (MR) analyses to detect causality. RESULTS: The metabolic signature comprised 81 metabolites, robustly correlated with a healthy lifestyle (r = 0.45, P = 4.2 × 10-15). The metabolic signature was inversely associated with RA risk (HR per standard deviation (SD) increment: 0.76; 95% CI: 0.70-0.83), and largely explained the protective effects of healthy lifestyle on RA with 64% (95% CI: 50.4-83.3) mediation proportion. 1- and 2-sample MR analyses also consistently showed the associations of genetically inferred per SD increment in metabolic signature with a reduction in RA risk (HR: 0.84; 95% CI: 0.75-0.94; and P = 0.002 and OR: 0.84; 95% CI: 0.73-0.97; and P = 0.02, respectively). CONCLUSIONS: Our findings implicate that the metabolic signature reflecting healthy lifestyle is a potential causal mediator in the development of RA, highlighting the importance of early lifestyle intervention and metabolic status tracking for precise prevention of RA.

Upscale production of (R)-mandelic acid with a stereospecific nitrilase in an aqueous system.

(R)-Mandelic acid (R-MA) is a key precursor for the synthesis of semi-synthetic penicillin, cephalosporin, anti-obesity drugs, antitumor agents, and chiral resolving agents for the resolution of racemic alcohols and amines. In this study, an enzymatic method for the large-scale production of R-MA by a stereospecific nitrilase in an aqueous system was developed. The nitrilase activity of the Escherichia coli BL21(DE3)/pET-Nit whole cells reached 138.6 U/g in a 20,000-L fermentor. Using recombinant E. coli cells as catalyst, 500 mM R,S-mandelonitrile (R,S-MN) was resolved into 426 mM (64.85 g/L) R-MA within 8 h, and the enantiomeric excess (ee) value of R-MA reached 99%. During the purification process, pure R-MA with a recovery rate of 78.8% was obtained after concentration and crystallization. This study paved the foundation for the upscale production of R-MA using E. coli whole cells as biocatalyst.

Dissipation Behavior of Three Fungicides during the Industrial Processing of PaeoniaeRadix Alba and Associated Processing Factors.

Before being administered as medicinal products, Chinese herbal medicines (CHMs) must be processed and decocted for human consumption. While the presence of pesticide residues in CHMs is a major concern, pesticide dissipation behavior during CHM processing has rarely been reported. In this study, the dissipation of three pesticide residues in the CHM Paeoniae Radix Alba (PRA) was investigated during each step of industrial processing. The boiling process was found to significantly reduce pesticide residues (61-89%), and the peeling process also contributed to pesticide degradation (29-68%). The high temperature (60 °C) during the drying process led to further pesticide degradation. The processing factors of all three pesticides after each processing step were less than one, and the processing factors for the overall process were lower than 0.027, indicating that industrial processing clearly reduced the amount of pesticide residues (97.3-99.4%). The findings provide guidance for the safe use of fungicides in CHMs and can help establish maximum residue limits for PRA to reduce human exposure to pesticides.

Significant improvement of the nitrilase activity by semi-rational protein engineering and its application in the production of iminodiacetic acid.

Iminodiacetic acid (IDA) is widely used as an intermediate in the manufacturing of chelating agents, glyphosate herbicides and surfactants. To improve activity and tolerance to the substrate for IDA production, Acidovorax facilis nitrilase was selected for further modification by the gene site saturation mutagenesis method. After screened by a two-step screening method, the best mutant (Mut-F168V/T201N/S192F/M191T/F192S) was selected. Compared to the wild-type nitrilase, Mut-F168V/T201N/S192F/M191T/F192S showed 136% improvement in specific activity. Co2+ stimulated nitrilase activity, whereas Cu2+, Zn2+ and Tween 80 showed a strong inhibitory effect. The Vmax and kcat of Mut-F168V/T201N/S192F/M191T/F192S were enhanced 1.23 and 1.23-fold, while the Km was decreased 1.53-fold. The yield of Mut-F168V/T201N/S192F/M191T/F192S with 453.2 mM of IDA reached 71.9% in 5 h when 630 mM iminodiacetonitrile was used as substrate. This study indicated that mutant nitrilase obtained in this study is promising in applications for the upscale production of IDAN.

Production of R-Mandelic Acid Using Nitrilase from Recombinant E. coli Cells Immobilized with Tris(Hydroxymethyl)Phosphine.

Recombinant Escherichia coli cells harboring nitrilase from Alcaligenes faecalis were immobilized using tris(hydroxymethyl)phosphine (THP) as the coupling agent. The optimal pH and temperature of the THP-immobilized cells were determined at pH 8.0 and 55 °C. The half-lives of THP-immobilized cells measured at 35, 40, and 50 °C were 1800, 965, and 163 h, respectively. The concentration of R-mandelic acid (R-MA) reached 358 mM after merely 1-h conversion by the immobilized cells with 500 mM R,S-mandelonitrile (R,S-MN), affording the highest productivity of 1307 g L-1 day-1 and the space-time productivity of 143.2 mmol L-1 h-1 g-1. The immobilized cells with granular shape were successfully recycled for 60 batches using 100 mM R,S-MN as substrate at 40 °C with 64% of relative activity, suggesting that the immobilized E. coli cells obtained in this study are promising for the production of R-MA.

Clinical comparison of the efficacy of three different bowel preparation methods on the infectious complications following transrectal ultrasonography-guided prostate biopsy in nursing practice.

AIMS AND OBJECTIVES: To assess the effects of three different bowel preparation methods on the incidence of infectious complications in patients who underwent transrectal ultrasonography-guided prostate biopsy. BACKGROUND: The standard bowel preparation protocol for prostate biopsy has not been established. DESIGN: A retrospective study in a single centre. METHODS: From January 2013-December 2015, the clinical records of 1,130 patients who underwent prostate biopsy were, respectively, reviewed. All the patients received metronidazole prophylaxis before biopsy. The patients were divided into three groups according to the bowel preparation methods: patients in Group A (n = 402) received only soapy enema; patients in Group B (n = 413) received polyethylene glycol; while patients in Group C (n = 315) received polyethylene glycol plus povidone-iodine enema. Infectious complications were classified as fever (>37.5°C), urinary tract infection and sepsis. The postoperative adverse events were also observed. RESULTS: The overall postbiopsy infectious complications were observed in 48 (4.25%) patients of all the cases, including 23 (5.72%) cases in Group A, 20 (4.84%) cases in Group B and five patients (1.59%) in Group C. There was significant difference among the groups (p = .018). In detail, these infectious complications included 22 (1.95%) cases of fever and 26 (2.30%) cases of urinary tract infection. No sepsis was observed among the total patients. The incidence of adverse events was 14.43% (58/402) occurred in Group A, 25.91% (107/413) in Group B and 26.67% (84/315) in Group C. The difference was statistically significant. CONCLUSIONS: Our study confirmed that combined preparation regimens of polyethylene glycol with povidone-iodine enema could significantly reduce the postbiopsy infection rate. Conventional soapy enema is associated with less adverse events. RELEVANCE TO CLINICAL PRACTICE: Findings of this study provide useful evidence-based information for healthcare professionals. The application of combined preparation regimens of polyethylene glycol with povidone-iodine enema resulted in better improvement in the prevention of postbiopsy infection.

Detection rate of prostate cancer following biopsy among the northern Han Chinese population: a single-center retrospective study of 1022 cases.

BACKGROUND: Prostate cancer is known to have ethnic and regional differences. The study aimed to clinically evaluate the detection rate of prostate cancer on transrectal ultrasonography (TRUS)-guided prostate biopsy and analyze its characteristics among the northern Han Chinese population at a single center. METHODS: Between October 2009 and September 2016, a total of 1027 Chinese men, who had undergone TRUS-guided prostate biopsy at Qingdao Municipal Hospital, were retrospectively analyzed. Prostate biopsies were performed in the case of an abnormally elevated serum PSA level, and/or abnormal digital rectal examination (DRE) findings, and/or suspicious prostatic imaging findings. RESULTS: Of the 1022 men enrolled in the analysis, 438 patients (42.8%) were diagnosed with prostate adenocarcinoma histologically. When serum PSA levels were divided into five subgroups (less than 4.0, 4.0 to 10.0, 10.0 to 20.0, 20.0 to 100.0, and ≥ 100.0 ng/ml), the detection rates of prostate cancer were 12.4, 15.9, 34.1, 66.2, and 93.8%, respectively. With serum PSA levels of 4.0 to 10.0 ng/ml, the cancer detection rates for a normal DRE and a suspect DRE finding were 13.5 and 58.2%, respectively. Accordingly, the cancer detection rates for a normal imaging and a suspect imaging finding were 13.5 and 58.2%, respectively. Besides, a large proportion of the patients were in the clinically advanced stage. CONCLUSIONS: The present study data reported a relatively higher prostate cancer detection rate of 42.8% and that the majority of the patients presented with clinically advanced prostate cancers within a local clinical urologic practice. An early detection and screening program for prostate cancer is of great need to reduce the burden from this disease among the northern Han Chinese population.

Downregulation of fatty acid synthase complex suppresses cell migration by targeting phosphor-AKT in bladder cancer.

The aim of the present study was to investigate the effect of fatty acid synthase complex (FASN) on the migration capacity of bladder transitional cell carcinoma (BTCC) cells and the involvement of matrix metalloproteinase­9 (MMP­9) via targeting of phospho­AKT (p­AKT). FASN­specific small­interfering RNA (FASN­siRNA) was used to inhibit FASN gene expression in the 5637 and 253J BTCC cell lines. The knockdown efficiency of FAM­conjugated FASN­siRNA was confirmed by fluorescence microscopy. The migratory abilities of BTCC cells were assessed using a Transwell assay. Furthermore, protein and mRNA expression of FASN, p­AKT, AKT, and migration­associated protein MMP­9 were detected by western blot analysis. Treatment with FASN inhibitor Cer and FASN­siRNA decreased the migratory capacity of bladder cancer cells and reduced the levels of p­AKT as well as the expression of MMP­9. These results indicated that FASN inhibition suppressed the migratory capacity of BTCC cells through suppressing AKT activation and consequently reducing MMP­9 expression. Targeting FASN may represent a promising novel therapeutic strategy for BTCC.

Complete mitochondrial genome sequence of a chronic obstructive pulmonary disease Rattus norvegicus C57CE strain.

We sequenced a chronic obstructive pulmonary disease Rattus norvegicus C57CE strain mitochondrial genome for the first time (GenBank Accession No. KM114607). Its mitogenome was 16,307 bp and coding 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes. This genome will provide definite information for genetic perspective into this disease.

Nitrilase-catalyzed conversion of (R,S)-mandelonitrile by immobilized recombinant Escherichia coli cells harboring nitrilase.

(R)-(-)-Mandelic acid (R-MA) is widely used both as a versatile intermediate for pharmaceuticals and a resolving agent in chiral resolution processes. In the current study, to improve the stability of operation, recombinant Escherichia coli cells expressing nitrilase from Alcaligenes faecalis were immobilized for the enantioselective hydrolysis of (R,S)-mandelonitrile to R-MA. Different immobilization methods including entrapment matrices, entrapment matrices cross-linked by cross-linking and polymerization agents, and direct cross-linking cells using glutaraldehyde (GA) or bionic silicon were investigated. To facilitate industrial solid-liquid separation, the direct cross-linking recombinant E. coli cells using diatomite/GA/polyethyleneimine with 135.95% relative activity compared with free cells was chosen using water as the reaction medium. The operational stability of the immobilized cells was obviously superior to that of free cells, without significant activity loss after 28 cycles of batch reaction and the successive production of R-MA could reach 1.88 M. Moreover, the immobilized cells showed good storage stability with about 52% relative activity after storing for 30 days at 4 °C. Therefore, the immobilized biocatalyst is very promising for upscale production of optically pure R-MA with high performance and low cost.

Improvement of Alcaligenes faecalis nitrilase by gene site saturation mutagenesis and its application in stereospecific biosynthesis of (R)-(-)-mandelic acid.

Nitrilases have recently received considerable attention as the biocatalysts for stereospecific production of carboxylic acids. To improve the activity, the nitrilase from Alcaligenes faecalis was selected for further modification by the gene site saturation mutagenesis method (GSSM), based on homology modeling and previous reports about mutations. After mutagenesis, the positive mutants were selected using a convenient two-step high-throughput screening method based on product formation and pH indicator combined with the HPLC method. After three rounds of GSSM, Mut3 (Gln196Ser/Ala284Ile) with the highest activity and ability of tolerance to the substrate was selected. As compared to the wild-type A. faecalis nitrilase, Mut3 showed 154% higher specific activity. Mut3 could retain 91.6% of its residual activity after incubation at pH 6.5 for 6 h. In a fed-batch reaction with 800 mM mandelonitrile as the substrate, the cumulative production of (R)-(-)-mandelic acid after 7.5 h of conversion reached 693 mM with an enantiomeric excess of 99%, and the space-time productivity of Mut3 was 21.50-fold higher than that of wild-type nitrilase. The Km, Vmax, and k(cat) of wild-type and Mut3 for mandelonitrile were 20.64 mM, 33.74 µmol mg(-1) min(-1), 24.45 s(-1), and 9.24 mM, 47.68 µmol mg(-1) min(-1), and 34.55 s(-1), respectively. A homology modeling and molecular docking study showed that the diameter of the catalytic tunnel of Mut3 became longer and that the tunnel volume was smaller. These structural changes are proposed to improve the hydrolytic activity and pH stability of Mut3. Mut3 has the potential for industrial applications in the upscale production of (R)-(-)-mandelic acid.

Biosynthesis of iminodiacetic acid from iminodiacetonitrile by immobilized recombinant Escherichia coli harboring nitrilase.

Iminodiacetic acid (IDA) is widely used as an intermediate in the manufacture of chelating agents, glyphosate herbicides and surfactants. In the current work, the fragment with the length of 1,110 bp encoding the Acidovorax facilis nitrilase was obtained. The recombinant nitrilase expressed in Escherichia coli BL21 (DE3) was successfully used in the production of IDA from iminodiacetonitrile. To improve the stability of operation, the recombinant cells were entrapped in polyvinyl alcohol (PVA) and sodium alginate (SA) copolymer. The maximum relative nitrilase activity with 98.1% was further observed at 1.0% SA, 8.0% PVA, 1.0% CaCl(2), and 5.0% wet cells, under conditions of 1.0% iminodiacetonitrile in distilled water and a temperature of 40°C, respectively. The entrapped cells facilitated easy separation and good recycling compared with free cells. Moreover, the immobilized cells showed good operation and storage stability. This report is the first to describe IDA preparation using immobilized recombinant E. coli harboring nitrilase.

Urinary trypsin inhibitor attenuates seawater-induced acute lung injury by influencing the activities of nuclear factor-ĸB and its related inflammatory mediators.

BACKGROUND: Few data are available on the role of neutrophil elastase (NE) and nuclear factor-κB (NF-κB) in the course of seawater drowning-induced acute lung injury (SWD-ALI), and there is no evidence on the value of giving urinary trypsin inhibitor (UTI) in the case of SWD-ALI. OBJECTIVE: To investigate the role of NF-κB and NE in the pathogenesis of SWD-ALI and whether UTI treatment can attenuate SWD-ALI in rabbits. METHODS: Rabbits were randomly assigned to control, seawater drowning, and UTI treatment groups. The rabbits in the control group only suffered from intubation, whereas the rabbits in the seawater drowning group and the UTI treatment group received arterial injection of normal saline without/with 50,000 U/kg body weight of UTI after instillation of seawater into an endotracheal catheter. The activities or contents of NF-κB, MPO, NE, TNF-α, and IL-10 in lung tissue were measured by nonradioactive EMSA, biochemical methods, and ELISA, respectively. RESULTS: After the seawater challenge, all of the rabbits demonstrated immediate drops in arterial PaO(2)/FiO(2) and pronounced pulmonary edema and inflammatory cell infiltration with evidence of an increase in the ratio of wet weight to dry weight, lung permeability index, lung injury scores, and the activities or contents of NF-κB, NE, MPO, TNF-α, and IL-10. UTI treatment markedly attenuated lung histopathological changes with evidence of a decrease in all of the parameters, except for upregulation of IL-10. Arterial PaO(2)/FiO(2) was significantly improved after 6 h of UTI treatment. CONCLUSION: These results suggest that NF-κB and NE play an important role in SWD-ALI. UTI protects against SWD-ALI, at least partly, through inhibition of the enhanced local activity of NF-κB, contents of TNF-α and NE, and infiltration of neutrophils and promotion of the level of IL-10.

[Differences between seawater- and freshwater-induced lung injuries].

OBJECTIVE: To investigate the differences between the lung injuries induced by seawater and freshwater drowning in a rabbit model. METHODS: Forty-two New Zealand rabbits were divided randomly into three groups: control group (C, n = 18), freshwater drowning group (F, n = 12), seawater drowning group (S, n = 12). The drowning model was established by pouring seawater or freshwater (2 ml/kg) into the respiratory tract through a tracheal catheter. Mean arterial pressure (MAP) and heart rate (HR) were monitored continually. Respiratory rate (RR), blood gas analysis and electrolyte contents of every rabbit were observed at different time. The lung wet to dry weight (W/D) ratio and lung permeability index (LPI) were calculated. The contents of malondialdehyde (MDA), myeloperoxidase (MPO) and superoxide dismutase (SOD) were measured by biochemical method. The expressions of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) were detected by enzyme linked immunosorbent assay (ELISA). At the same time, the changes in pathology were studied with by hematoxylin eosin (HE) staining, and lung pathologic score (LPS) was calculated. RESULTS: There was no significant difference in blood electrolyte contents and HR among the three groups (all P > 0.05). In freshwater drowning, there was a temporary increase of MAP at 5 minutes, RR increased immediately, and partial pressure of carbon dioxide in artery (PaCO(2)) and base excess (BE) were persistently decreased (P < 0.05 or P < 0.01). Oxygenation index (PaO(2)/FiO(2)) fell to (297.8+/-81.3) mm Hg (1 mm Hg = 0.133 kPa) at 0.5 hour, then elevated to over 300 mm Hg rapidly, and then reverted to initiative level in around 2 hours. There were several edematous and petechial areas on the dependent region of the lung. Alveolar collapse and parenchymal congestion were the main pathological features. W/D ratio and LPI showed no remarkable change. In lung tissue, the level of LPS, MPO, MDA, TNF-alpha and IL-1 beta had a significant increase, while SOD had a significant decrease (P < 0.05 or P <0.01). In S group, respiratory symptoms were more serious; edematous and congestive areas of the lung were more extensive, and the dependent region showed hepatization changes. W/D ratio and LPI consisted of elevated significantly (all P < 0.01). The pathological characteristics were massive inflammatory cell infiltration and more serious alveolar edema. Compared with F group, the extent of up- or down-regulation of RR, MAP, PaO(2)/FiO(2), PaCO(2), BE, inflammatory media and cytokines in S group was more prominent and steady, and S 6-hour group had a higher pathological score than S 3-hour group and F group (P < 0.05 or P < 0.01). CONCLUSION: Seawater and freshwater drowning could not only injure pulmonary parenchymal cells directly, but also induce acute inflammatory reaction. Lung injury induced by seawater is severer than that by freshwater.