Insight into the Role of an α-Helix Cluster in Protoporphyrinogen IX Oxidase.

Protoporphyrinogen IX oxidase (PPO) is the last common enzyme in chlorophyll and heme biosynthesis pathways. In humans, point mutations on PPO are responsible for the dominantly inherited disorder disease variegate porphyria (VP). It is found that several VP-causing mutation sites are located on an α-helix cluster (consisting of α-5, α-6, and α-7 helix, named the G169 helix cluster) of human PPO, although these mutation sites are outside the active site of the human PPO. In this work, we investigated the role of the G169 helix cluster via site-directed mutagenesis, enzymatic kinetics, and computational studies. Kinetic studies showed that mutations on the G169 helix cluster affect the activity of PPO. The MD simulation showed that mutations on the G169 helix cluster reduced the activity of PPO by affecting the proper orientation of substrate protoporphyrinogen within the active site of PPO and possibly the dipole moment of the G169 helix cluster. Moreover, the mutation abolished the interaction between the mutated site and other residues, thus affecting the secondary structure and hydrogen bond interactions within the G169 helix cluster. These results indicated that the integrity of the G169 helix cluster is important for the stabilization of protoporphyrinogen within the active site of PPO to facilitate the interaction between protoporphyrinogen and cofactor FAD and provide a proper electrostatic environment for the activity of PPO. Our result provides new insight into understanding the relationship between the structure and function of PPO.

Risk Factors for Breast Cancer-Related Lymphedema: An Umbrella Review.

BACKGROUND: Identification of risk factors facilitates the prevention of breast cancer-related lymphedema (BCRL). Several published systematic reviews have already addressed the risk factors for BCRL. This study aimed to systematically identify potential risk factors for BCRL and evaluate the quality of evidence. METHODS: The study followed methodologic guidance from the Joanna Briggs Institute, and the Cochrane Handbook. The following electronic databases were systematically searched from inception to 15 November 2022: PubMed, Embase, CINAHL, Web of Science, Scopus, CNKI, SinoMed, Wanfang, JBI Database, Cochrane Database, ProQuest, and PROSPERO. Two authors independently screened studies, extracted data, and assessed methodologic quality using AMSTAR2, risk of bias using ROBIS, and evidence quality using GRADE. The study evaluated overlap, assessed the small-study effect, and calculated the I2 statistic and Egger's P value as needed. RESULTS: The study included 14 publications comprising 10 meta-analyses and 4 systematic reviews. The authors identified 39 factors and 30 unique meta-analyses. In the study, 13 innate personal trait-related risk factors, such as higher body mass index (BMI) and axillary lymph nodes dissection, showed statistically significant associations with BCRL incidence. Breast reconstruction was found to be a protective factor. The methodologic quality was low or critically low. The majority of the systematic reviews and/or meta-analyses were rated as having a high risk of bias. Evidence quality was low for 22 associations and moderate for 8 associations. CONCLUSIONS: The currently identified risk factors for BCRL all are innate personal trait-related factors. Future well-designed studies and robust meta-analyses are needed to explore potential associations between behavioral-, interpersonal-, and environmental-related factors and BCRL, as well as the role of genetic variations and pathophysiologic factors.

Contemporaneous Symptom Networks of Breast Cancer-Related Upper Limb Lymphedema: A Network Analysis.

BACKGROUND: Upper limb lymphedema (ULL) is a common and deliberating complication for breast cancer survivors (BCSs). Breast cancer survivors with ULL reported a wide range of symptoms. However, little is known about symptom patterns and interrelationships among them. This study was designed to explore symptom clusters and construct symptom networks of ULL-related symptoms among BCSs and to identify the core symptoms. METHODS: This study is a secondary data analysis using datasets from three cross-sectional studies of BCSs in China. A total of 341 participants with maximum interlimb circumference ≥2 cm and complete responses in Part I of the Breast Cancer and Lymphedema Symptom Experience Index were included. Symptom clusters were identified through principal component analysis, and multiple linear regression analysis was employed to explore factors associated with severity of overall ULL-related symptoms. A contemporaneous network with 20 frequently reported symptoms were constructed after controlling for covariates. RESULTS: Three symptom clusters, including lymph stasis symptom cluster, nerve symptom cluster, and movement limitation symptom cluster, were identified. Postsurgery time, axillary lymph node dissection, and radiotherapy were associated with the severity of ULL-related symptoms. Tightness (rs = 1.379; rscov = 1.097), tingling (rs = 1.264; rscov = 0.925), and firmness (rs = 1.170; rscov = 0.923) were the most central symptoms in both networks with and without covariates. CONCLUSIONS: Breast cancer survivors with ULL experienced severe symptom burden. Tightness, tingling, and firmness were core symptoms of ULL among BCSs. Our findings demonstrated that the assessment and targeted intervention of specific core symptoms might help to relive effectively the burden of ULL-related symptom among BCSs.

Efficacy of pramipexole on quality of life in patients with Parkinson's disease: a systematic review and meta-analysis.

BACKGROUND: Quality of life (QoL) in patients with Parkinson's disease (PD) is increasingly used as an efficacy outcome in clinical studies of PD to evaluate the impact of treatment from the patient's perspective. Studies demonstrating the treatment effect of pramipexole on QoL remain inconclusive. This study aims to evaluate the effect of pramipexole on QoL in patients with PD by conducting a systematic review and meta-analysis of existing clinical trials. METHODS: A systematic literature search of PubMed, Embase and the Cochrane Library was performed from inception to 30 April 2022 to identify randomised, placebo-controlled trials of patients with idiopathic PD receiving pramipexole, who reported a change from baseline in their QoL as measured by the 39-item Parkinson's Disease Questionnaire (PDQ-39). Risk of bias was independently assessed by two reviewers using the Cochrane Collaboration's tool for bias assessment. RESULTS: Of 80 eligible articles screened, six trials consisting of at least 2000 patients with early or advanced PD were included. From the synthesis of all six selected trials, a significant mean change from baseline in the PDQ-39 total score of -2.49 (95% CI, -3.43 to -1.54; p < 0.0001) was observed with pramipexole compared with placebo. A trend toward improvement in QoL was consistently observed among patients who received optimal doses of pramipexole (≥ 80% of the study population on 1.5 mg dosage), regardless of disease severity (advanced versus early) or baseline QoL levels. CONCLUSION: This meta-analysis provides evidence for the potential treatment benefit of pramipexole in improving QoL in patients with PD.

Parkinson's disease is a chronic, progressive nervous system disorder with no known cure. Patients may experience a number of symptoms including stiffness, slowness, and uncontrollable muscle movements, all of which impact their quality of life. Most clinical studies in Parkinson's disease measure the effect of treatment on improving symptoms, but other aspects such as quality of life are often overlooked. It is important to include quality of life measures in clinical studies of Parkinson's disease, such as the 39-item Parkinson's disease questionnaire (PDQ-39), to understand the impact of treatment from patients' perspectives. Pramipexole is a dopamine agonist that is well-tolerated and effective at treating Parkinson's disease symptoms. However, studies examining its effect on quality of life are inconclusive. This meta-analysis of existing clinical trials therefore aimed to evaluate the effect of pramipexole on quality of life in patients with Parkinson's disease. Six trials consisting of at least 2000 patients with early or advanced Parkinson's disease receiving treatment with pramipexole were included in this meta-analysis. Analysis of these six trials found a significant improvement in PDQ-39 total score with pramipexole compared with placebo. This meta-analysis provides new evidence for the potential treatment benefit of pramipexole in improving quality of life in patients with Parkinson's disease.

The hydrogen bonding network involved Arg59 in human protoporphyrinogen IX oxidase is essential for enzyme activity.

Protoporphyrinogen IX oxidase (PPO) is the last common enzyme in chlorophyll and heme biosynthesis pathways. In human, point mutations on PPO are responsible for the dominantly inherited disorder disease, Variegate Porphyria (VP). Of the VP-causing mutation site, the Arg59 is by far the most prevalent VP mutation residue identified. Multiple sequences alignment of PPOs shows that the Arg59 of human PPO (hPPO) is not conserved, and experiments have shown that the equivalent residues in PPO from various species are essential for enzymatic activity. In this work, it was proposed that the Arg59 performs its function by forming a hydrogen-bonding (HB) network around it in hPPO, and we investigated the role of the HB network via site-directed mutagenesis, enzymatic kinetics and computational studies. We found the integrity of the HB network around Arg59 is important for enzyme activity. The HB network maintains the substrate binding chamber by holding the side chain of Arg59, while it stabilizes the micro-environment of the isoalloxazine ring of FAD, which is favorable for the substrate-FAD interaction. Our result provides a new insight to understanding the relationship between the structure and function for hPPO that non-conserved residues can form a conserved element to maintain the function of protein.

Nucleostemin promotes hepatocellular carcinoma by regulating the function of STAT3.

Hepatocellular carcinoma (HCC) is the most common primary malignant tumor in the liver and the second leading cause of cancer-related death worldwide. The collaborative function between Nucleostemin (NS) and STAT3 has been reported but not well studied in HCC. Here, we found a significant correlation between NS expression and STAT3 phosphorylation, not only in HCC cancers but also in HCC tissues. Patients with high expression of both NS and p-STAT3 show a very poor survival rate. High expression of both NS and p-STAT3 is also associated with tumor size and microvascular invasion. Knocking down the expression of NS greatly reduces the phosphorylation of STAT3. Conversely, overexpression of NS significantly promotes STAT3 phosphorylation. NS and p-STAT3 are located in the nucleus and physiologically interact with each other. Furthermore, NS greatly enhances cell migration and invasion by promoting the epithelial-mesenchymal transition (EMT). NS also supports cell proliferation and colony formation. The importance of NS in HCC was further demonstrated by evaluating tumor formation in vivo. Therefore, we demonstrate a critical collaborative function between NS and STAT3 in HCC, providing an invaluable insight into the mechanism of HCC. The concomitant expression of NS and p-STAT3 might be a potential prognostic indicator and therapeutic target in patients with HCC.

Driving GABAergic neurons optogenetically improves learning, reduces amyloid load and enhances autophagy in a mouse model of Alzheimer's disease.

The changes of local field potentials (LFP, mainly gamma rhythm and theta rhythm) in the brain are closely related to learning and memory formation. Reduced gamma rhythm (20-50 Hz) and theta rhythm (4-10 Hz) has been observed in the progression of Alzheimer's disease (AD), but it is not clear whether it is related to cognition in AD. Here, we investigated behaviorally driven gamma rhythm and theta rhythm in APP/PS1 mice, and optogenetically stimulated GABAergic neurons in the brain to better understand the relationship between the changes of LFP, cognition, and cellular pathologies. Optogenetically driving GABAergic neurons rescued memory formation in a water maze task and normalized theta and gamma rhythm in the EEG. Furthermore, the optogenetic stimulation alleviated neuroinflammation and levels of amyloid-ß (Aß)1-42 fragments, and induced autophagy. GABA blockers also reversed the normalization of theta and gamma rhythms in the brain by optogenetic stimulation. The results demonstrate that stimulation of GABAergic interneurons not only rescues LFP rhythms and memory formation, but furthermore activates autophagy and reduces neuroinflammation, which have beneficial additional effects such as clearing amyloid. This is a proof of concept for a novel therapeutic approach to AD treatment.

Determining multi-pesticide residues in teas by dispersive solid-phase extraction combined with speed-regulated directly suspended droplet microextraction followed by gas chromatography-tandem mass spectrometry.

In this study, an effective speed-regulated directly suspended droplet microextraction method was developed to condense pesticide residues from teas through dispersive solid-phase extraction prior to analysis by gas chromatography with tandem mass spectrometry. The extractant was intentionally dispersed into the sample solution in the form of globules through high-speed agitation. This procedure increases the contact area between the binary phases and shortens the distribution equilibrium time. The fine globules reassembled by decelerating stirring speed, the extractant could be taken out for gas chromatography with tandem mass spectrometry. Recovery studies were performed under optimized extraction conditions by using matrix blanks fortified with pesticides at three concentrations (10, 50, and 100 µg/kg). Over 87% of the recoveries for the analytes in four tea matrices were acceptable given their recovery ranges of 70-120% and relative standard deviations of ≤20%. The limits of quantification of most pesticides were lower than 10 µg/kg and thus satisfied the requirements for maximum residue levels prescribed by the European Community. A total of 38 tea samples from local markets were analyzed by using the proposed method. Results showed that chlorpyrifos was the most frequently detected pesticide in teas. The method is a potential choice for the routine monitoring of pesticide residues in complex matrices.

Determination of multiple pesticide residues in teas by gas chromatography with accurate time-of-flight mass spectrometry.

In this work, gas chromatography tandem with electron ionization and full-scan high-resolution mass spectrometry with a time-of-flight mass analyzer was evaluated for analyzing pesticide residues in teas. The relevant aspects for mass spectrometry analysis, including the resolution and mass accuracy, acquisition rate, temperature of ion source, were investigated. Under acquisition condition in 2-GHz extended dynamic range mode, accurate mass spectral library including 184 gas chromatography detectable pesticides was established and retrieval parameters were optimized. The mass spectra were consistent over a wide concentration range (three orders) with good match values to those of NIST (EI-quadrupole). The methodology was verified by the validation of 184 pesticides in four tea matrices. A wide linear range (1-1000 µg/kg) was obtained for most compounds in four matrices. Limit of detection, limit of quantification, and limit of identification values acquired in this study could satisfy the requirements for maximum residue levels prescribed by the European Community. Recovery studies were performed at three concentrations (10, 50, and 100 µg/kg). Most of the analytes were recovered at an acceptable range of 70-120% with relative standard deviations ≤ 20% in four matrices. The potential extension of qualitative screening scope makes gas chromatography tandem with electron ionization and mass spectrometry with a time-of-flight mass analyzer a more powerful tool compared with gas chromatography with tandem mass spectrometry.

Effects of Leptin on Na+/Ca2+ Exchanger in PC12 Cells.

BACKGROUND/AIMS: Alzheimer's disease (AD) is known to be related to alterations in neuronal intracellular calcium activity ([Ca2+]i). The present study revealed the distinct role of leptin in Na+/Ca2+-exchanger activity. METHODS: [Ca2+]i was determined utilizing Fura-2 fluorescence. The activity of NCX was measured by removal of extracellular Na+ in the presence of external Ca2+. Na+/Ca2+-exchanger activity was further quantified from whole cell currents following removal of extracellular Na+. Na+/Ca2+-exchanger isoform NCX1 transcript levels and protein abundance were quantified by RT-PCR and Western blotting, respectively. RESULTS: Exposure of PC12 cells to 30 µM amyloid (Aß42) increased [Ca2+]i, an effect significantly blunted by 6 hours incubation with leptin before Aß42 treatment. Moreover, leptin treatment significantly increased Na+/Ca2+-exchanger mediated Ca+ transport and current, NCX1 transcript level as well as NCX1 membrane protein abundance. CONCLUSION: We show that leptin blunts Aß42-evoked [Ca2+]i increase by increasing expression and activity of Na+/Ca2+-exchanger NCX1.

Leptin Regulates Tau Phosphorylation through Wnt Signaling Pathway in PC12 Cells.

BACKGROUND/AIMS: Leptin, an adipocytokine produced endogenously in the brain, is decreased in Alzheimer's disease(AD) and has also been shown to reduce Aß levels in vitro and in vivo. Sets of evidence show that leptin reduces Aß production and tau phosphorylation in neuronal cells and transgenic mice models of AD. Herein, we investigated the signaling pathway activated by leptin, to better understand its mechanism of action. METHODS: Western blotting was performed to assess the levels of phosphor-tau and Bax, RT-PCR to check the mRNA level of Bax. RESULTS: Leptin treatment significantly blunted Aß-evoked tau phosphorylation and Bax levels, effects of which could be reversed by antagonist of Wnt signaling. CONCLUSION: The data indicate that Leptin may provide a novel therapeutic approach to AD treatment via wnt signaling.

Fluorescent visualization of Src by using dasatinib-BODIPY.

Many biological experiments are not compatible with the use of immunofluorescence, genetically encoded fluorescent tags, or FRET-based reporters. Conjugation of existing kinase inhibitors to cell-permeable fluorophores can provide a generalized approach to develop fluorescent probes of intracellular kinases. Here, we report the development of a small molecule probe of Src through conjugation of BODIPY to two well-established dual Src-Abl kinase inhibitors, dasatinib and saracatinib. We show that this approach is not successful for saracatinib but that dasatinib-BODIPY largely retains the biological activity of its parent compound and can be used to monitor the presence of Src kinase in individual cells by flow cytometry. It can also be used to track the localization of Src by fixed and live-cell fluorescence microscopy. This strategy could enable generation of additional kinase-specific probes useful in systems not amenable to genetic manipulation or could be used together with fluorescent proteins to enable a multiplexed assay readout.

Metformin reverses multidrug resistance and epithelial-mesenchymal transition (EMT) via activating AMP-activated protein kinase (AMPK) in human breast cancer cells.

Breast cancer is the most frequently diagnosed tumor type and the primary leading cause of cancer deaths in women worldwide and multidrug resistance is the major obstacle for breast cancer treatment improvement. Emerging evidence suggests that metformin, the most widely used antidiabetic drug, resensitizes and cooperates with some anticancer drugs to exert anticancer effect. However, there are no data regarding the reversal effect of metformin on chemoresistance in breast cancer. In the present study, we investigated the resistance reversal effect of metformin on acquired multidrug-resistant breast cancer cells MCF-7/5-Fu derived from MCF-7 breast cancer cells and innate multidrug-resistant MDA-MB-231 breast cancer cells, and we found that metformin resensitized MCF7/5-FU and MDA-MB-231 to 5-fluorouracil (5-FU), adriamycin, and paclitaxel. We also observed that metformin reversed epithelial-mesenchymal transition (EMT) phenotype and decreased the invasive capacity of MCF7/5-FU and MDA-MB-231 cells. However, there were no significant changes upon metformin-treated MCF7 cells. Moreover, we found metformin treatment activated AMPK signal pathway in MCF7/5-FU and MDA-MB-231 cells and compound C, the AMPK inhibitor, could partly abolish the resensitization and EMT reversal effect of metformin. To the best of our knowledge, we are the first to report that metformin can resensitize multidrug-resistant breast cancer cells due to activating AMPK signal pathway. Our study will help elucidate the mechanism of chemoresistance and establish new strategies of chemotherapy for human breast cancer.

Meta-analysis of changes in thiol-disulfide homeostasis during preeclampsia.

The present study systematically assessed alterations in thiol-disulfide homeostasis among women with preeclampsia (PE) through meta-analysis. This was conducted as such changes are believed to be associated with the oxidative stress underlying this condition. A comprehensive search of Medline, Web of Science, and Embase databases was conducted from their inception until 22 March 2023, to identify studies comparing levels of native thiol, total thiol, and disulfide between pregnant women with PE and those without PE. Results were pooled using a random-effects model to account for study heterogeneity. The analysis included a total of 631 women diagnosed with PE and 668 healthy pregnant women, encompassing 13 case-control studies and 1 prospective study. Pooled outcomes revealed that women with PE had significantly lower blood levels of native thiol, (mean difference [MD] -51.42 umol/L; 95% confidence interval [CI] -79.75 to -23.10 umol/L; P < 0.001; I2 = 0% and total thiol (MD -65.56 umol/L; 95% CI -104.97 to -26.15 umol/L; P = 0.001; I2 = 0%) compared to the control group. In contrast, no significant difference was observed in blood disulfide levels between the two groups (MD -1.10 umol/L; 95% CI -4.41 to -2.21 umol/L; P = 0.51; I2 = 0%). Subgroup analyses indicated that the results were consistent across studies matched by gestational age and body mass index, as well as those with varying quality scores (P for subgroup differences all > 0.05). In conclusion, women with PE are associated with significantly reduced blood levels of native and total thiols but show no change in blood disulfide levels, suggesting a state of reduced antioxidants in PE.

A Comprehensive Study of Heterogeneous Mismatch Repair Expression in Solid Tumors Reveals Different Immunohistochemical Patterns and Distinct Genetic Mechanisms.

OBJECTIVE: Immunohistochemistry is routinely performed to detect mismatch repair deficiency in solid tumors. Heterogeneous MMR expression (MMR-het) has been reported occasionally but not systemically studied. METHODS: In this study, we depicted MMR-het patterns of 40 tumors of different anatomical sites and analyzed MMR genetic alterations and tumor mutational burdens (TMB) through comprehensive genomic profiling. RESULTS: The MMR-het patterns were classified into 4 subgroups: "single-loss" (3 cases), "MLH1/PMS2 double-loss" (16 cases), "MSH2/MSH6 double-loss" (8 cases), and "triple/tetra-loss" (13 cases). Seventeen MMR-het cases exhibited histological heterogeneity, in which MMR protein loss was generally confined to either poorly differentiated or well-differentiated tumor areas. All "single-loss" tumors had MMR somatic mutations and coexisting POLE exonuclease domain mutations. "MLH1/PMS2 double-loss" tumors unexceptionally harbored MLH1 hypermethylation without MMR germline mutations. In the "MSH2/MSH6 double-loss" subgroup, 4 cases had MSH2/MSH6 germline mutations, while another 4 cases had multiple MSH2/MSH6 somatic mutations. Additional POLE exonuclease domain mutations were identified in 2 cases. Tumors in the "triple/tetra-loss" subgroup generally had MLH1 abnormalities (8 MLH1 hypermethylation, 4 MLH1 germline mutation, 1 MLH1 double somatic mutations), and coexistent somatic mutations on MSH2/MSH6 . Thirty-one cases (83.8%) were TMB-H, and all POLE -mutated cases exhibited ultra-high TMB (111.4 to 524.2 mut/Mb). CONCLUSION: Our findings highlighted the importance of accurately interpreting heterogeneous MMR protein staining patterns for developing a more efficient personalized genetic investigation strategy.

Predict nutrition-related adverse outcomes in head and neck cancer patients undergoing radiotherapy: A systematic review.

BACKGROUND: Acute nutrition-related adverse outcomes are common in head and neck cancer patients undergoing radiotherapy. Predictive models can assist in identifying high-risk patients to enable targeted intervention. We aimed to systematically evaluate predictive models for predicting severe acute nutritional symptoms, insufficient intake, tube feeding, sarcopenia, and weight loss. METHODS: We searched PubMed, Web of Science, EBSCO, Embase, WanFang, CNKI, and SinoMed. We selected studies developing predictive models for the aforementioned outcomes. Data were extracted using a predefined checklist. Risk of bias and applicability assessment were assessed using the Prediction model Risk of Bias Assessment Tool. A narrative synthesis was conducted to summarize the model characteristics, risk of bias, and performance. RESULTS: A total of 2941 studies were retrieved and 19 were included. Study outcome measure were different symptoms (n = 11), weight loss (n = 5), tube feeding (n = 3), and symptom or tube feeding (n = 1). Predictive factors mainly encompassed sociodemographic data, disease-related data, and treatment-related data. Seventeen studies reported area under the curve or C-index values ranging from 0.610 to 0.96, indicating moderate to good predictive performance. However, candidate predictors were incomplete, outcome measures were diverse, and the risk of bias was high. Most of them used traditional model development methods, and only two used machine learning. CONCLUSIONS: Most current models showed moderate to good predictive performance. However, predictors are incomplete, outcome are inconsistent, and the risk of bias is high. Clinicians could carefully select the models with better model performance from the available models according to their actual conditions. Future research should include comprehensive and modifiable indicators and prioritize well-designed and reported studies for model development.

Specific Activation of Dopamine Receptor D1 Expressing Neurons in the PrL Alleviates CSDS-Induced Anxiety-Like Behavior Comorbidity with Postoperative Hyperalgesia in Male Mice.

Postoperative pain is a type of pain that occurs in clinical patients after surgery. Among the factors influencing the transition from acute postoperative pain to chronic postoperative pain, chronic stress has received much attention in recent years. Here, we investigated the role of dopamine receptor D1/D2 expressing pyramidal neurons in the prelimbic cortex (PrL) in modulating chronic social defeat stress (CSDS)-induced anxiety-like behavior comorbidity with postoperative hyperalgesia in male mice. Our results showed that preoperative CSDS induced anxiety-like behavior and significantly prolonged postoperative pain caused by plantar incision, but did not affect plantar wound recovery and inflammation. Reduced activation of dopamine receptor D1 or D2 expressing neurons in the PrL is a remarkable feature of male mice after CSDS, and chronic inhibition of dopamine receptor D1 or D2 expressing neurons in the PrL induced anxiety-like behavior and persistent postoperative pain. Further studies found that activation of D1 expressing but not D2 expressing neurons in the PrL ameliorated CSDS-induced anxiety-like behavior and postoperative hyperalgesia. Our results suggest that dopamine receptor D1 expressing neurons in the PrL play a crucial role in CSDS-induced anxiety-like behavior comorbidity with postoperative hyperalgesia in male mice.

Isolation, structural elucidation and biosynthetic pathway of bioactive prenyl quinone compounds from Panus lecomtei based on untargeted metabolomics combined with molecular networking.

Panus lecomtei is a relatively unfamiliar and undeveloped mushroom. This study generated ethyl acetate extracts of P. lecomtei intracellular (I), extracellular (E) and total fermentation broth (T). Both E and T extracts demonstrated antioxidant and antibacterial activities at 100 to 200 µg/mL. The composition differences of metabolites of these extracts were further studied based on comparative metabolomics by LS/MS and molecular network analysis. The results revealed that there were over 2000 significantly distinct metabolites among the three extracts, with abundant prenyl quinone compounds. Furthermore, the molecular network clarified the conversion relationship of P. lecomtei metabolites. Seven known prenyl quinone derivatives (1-7) were isolated from the E extract. Among them, compound 3 displayed excellent antioxidant activity and modest antibacterial activity. Compound 5 was discovered in fungi for the first time. Finally, a potential biosynthetic route for prenyl quinone in P. lecomtei was suggested.

Functional elucidation of the EIF4A3-circR-4225-miR-507-TNFSF11 regulatory axis in LUAD and its role in tumor progression.

Lung adenocarcinoma (LUAD) is the most common subtypes of NSCLC. However, the therapeutic effects for LUAD are unsatisfactory at current stage, so it is important to find new molecular targets and therapeutic strategies. circRNAs can regulate the expression of target genes by binding to microRNAs (miRNAs) to form competitive endogenous RNAs (ceRNAs). Therefore, we investigated the functions of circR-4225 in the tumor progression of LUAD and its molecular mechanism in this paper. circR-4225 is up-regulated in LUAD tissues. EIF4A3, a member of the eukaryotic translation initiation factor 4A (EIF4A) family, promotes the expression of circR-4225. circR-4225 acts as a molecular sponge to down-regulate miR-507, which promotes the up-regulation of the expression of its target gene-tumor necrosis factor superfamily member 11 (TNFSF11). Knockdown of circR-4225 in the LUAD cell lines can inhibit cell proliferation and viability, and promote apoptosis of the LUAD cell lines, which can be reverted by inhibiting miR-507 or overexpressing TNFSF11. To sum it up, this study demonstrated that circR-4225 was significantly up-regulated in LUAD tissues, and circR-4225 promoted LUAD progression by sponging miR-507 and up-regulating TNFSF11. This study can provide new molecular targets for early diagnosis and treatment of LUAD.

Enhanced production of N-acetyl-D-neuraminic acid by multi-approach whole-cell biocatalyst.

N-Acetyl-D-neuraminic acid (Neu5Ac) has attracted considerable interest due to its promising potential applications in medicine. Significant efforts have been made in whole-cell biocatalyst for Neu5Ac production, but the processes often result in suboptimal performance due to poor expression of enzymes, imbalances of pathway components, disturbance of competing pathways, and barriers of mass transport. In this study, we engineered Escherichia coli strains capable of producing Neu5Ac by assembling a two-step heterologous pathway consisting of N-acetyl-D-glucosamine 2-epimerase (AGE) and Neu5Ac aldolase (NanA). Multiple approaches were used to improve the efficiency of the engineered pathway and process for enhanced Neu5Ac production. Firstly, we identified that NanA was the rate-controlling enzyme in this pathway. With increased expression of NanA, a ninefold increase in Neu5Ac production (65 mM) was observed. Secondly, knocking out nanTEK genes blocked Neu5Ac uptake and the competing pathway, which kept the reactions to the synthetic direction as the final product went outside of the cells and enhanced the Neu5Ac production by threefold, resulting in 173.8 mM of Neu5Ac. Thirdly, we improved the performance of the system by promoting substrate transport and optimizing concentrations of substrates. An overall whole-cell biocatalytic process was developed and a maximum titer of 240 mM Neu5Ac (74.2 g/L) was achieved, with productivity of 6.2 g Neu5Ac/L/h and conversion yield of 40 % from GlcNAc. The engineered strain could be reused for at least five cycles with a productivity of >6 g/L/h. It is a cost-effective process for Neu5Ac production with potential applications in large-scale industrial production.