Distinct characteristics of the gut virome in patients with osteoarthritis and gouty arthritis.

BACKGROUND/PURPOSE(S): The gut microbiota and its metabolites play crucial roles in pathogenesis of arthritis, highlighting gut microbiota as a promising avenue for modulating autoimmunity. However, the characterization of the gut virome in arthritis patients, including osteoarthritis (OA) and gouty arthritis (GA), requires further investigation. METHODS: We employed virus-like particle (VLP)-based metagenomic sequencing to analyze gut viral community in 20 OA patients, 26 GA patients, and 31 healthy controls, encompassing a total of 77 fecal samples. RESULTS: Our analysis generated 6819 vOTUs, with a considerable proportion of viral genomes differing from existing catalogs. The gut virome in OA and GA patients differed significantly from healthy controls, showing variations in diversity and viral family abundances. We identified 157 OA-associated and 94 GA-associated vOTUs, achieving high accuracy in patient-control discrimination with random forest models. OA-associated viruses were predicted to infect pro-inflammatory bacteria or bacteria associated with immunoglobulin A production, while GA-associated viruses were linked to Bacteroidaceae or Lachnospiraceae phages. Furthermore, several viral functional orthologs displayed significant differences in frequency between OA-enriched and GA-enriched vOTUs, suggesting potential functional roles of these viruses. Additionally, we trained classification models based on gut viral signatures to effectively discriminate OA or GA patients from healthy controls, yielding AUC values up to 0.97, indicating the clinical utility of the gut virome in diagnosing OA or GA. CONCLUSION: Our study highlights distinctive alterations in viral diversity and taxonomy within gut virome of OA and GA patients, offering insights into arthritis etiology and potential treatment and prevention strategies.

Excess copper induced proteomic changes in the marine brown algae Sargassum fusiforme.

Copper (Cu) is an essential micronutrient for algal growth and development; however, it is also generally considered to be one of the most toxic metals when present at higher levels. Seaweeds are often exposed to low concentrations of metals, including Cu, for long time periods. In cases of ocean outfall, they may even be abruptly exposed to high levels of metals. The physiological processes that are active under Cu stress are largely unknown. In this study, the brown macroalga Sargassum fusiforme was cultured in fresh seawater at final Cu concentrations of 0, 4, 8, 24 and 47 µM. The Cu(2+) concentration and chlorophyll autofluorescence were measured to establish the toxic effects of Cu on this economically important seaweed. The accumulation of Cu by S. fusiforme was also dependent upon the external Cu concentration. Algal growth displayed a general decline with increasing media Cu concentrations, indicating that S. fusiforme was able to tolerate Cu stress at low concentrations, while it was negatively impacted at high concentrations. The term "acute stress" was employed to indicate exposure to high Cu concentrations for 1 day in this study. On the other hand, "chronic stress" was defined as exposure to lower sub-lethal Cu concentrations for 7 days. Proteins were extracted from control and Cu-treated S. fusiforme samples and separated by two-dimensional gel electrophoresis. Distinct patterns of protein expression in the acute and chronic stress conditions were observed. Proteins related to energy metabolism and photosynthesis were reduced significantly, whereas those related to carbohydrate metabolism, protein destination, RNA degradation and signaling regulation were induced in S. fusiforme in response to acute copper stress. Energy metabolism-related proteins were significantly induced by chronic Cu stress. Proteins from other functional groups, such as those related to membranes and transport, were present in minor quantities. These results suggest that S. fusiforme is sensitive to excess Cu, regardless of the presence of acute or chronic stress. We discuss the possible function of these identified proteins, taking into consideration the information available from other plant models.

Folate intake, serum folate levels, and prostate cancer risk: a meta-analysis of prospective studies.

BACKGROUND: Studies have reported inconsistent results concerning the existence of associations of folate intake and serum folate levels with prostate cancer risk. This study sought to summarise the evidence regarding these relationships using a dose-response meta-analysis approach. METHODS: In January 2014, we performed electronic searches of PubMed, Embase, and the Cochrane Library to identify studies examining the effect of folate on the incidence of prostate cancer. Only prospective studies that reported effect estimates with 95% confidence intervals (CIs) of the incidence of prostate cancer for more than 2 categories of folate were included. RESULTS: Overall, we included 10 prospective studies reporting data on 202,517 individuals. High dietary folate intake had little or no effect on prostate cancer risk (risk ratio [RR] = 1.02; 95% CI = 0.95-1.09; P = 0.598). The dose-response meta-analysis suggested that a 100 µg per day increase in dietary folate intake has no significant effect on the risk of prostate cancer (RR = 1.01; 95% CI = 0.99-1.02; P = 0.433). However, high serum folate levels were associated with an increased risk of prostate cancer (RR = 1.21; 95% CI = 1.05-1.39; P = 0.008). The dose-response meta-analysis indicated that a 5 nmol/L increment of serum folate levels was also associated with an increased risk of prostate cancer (RR = 1.04; 95% CI = 1.00-1.07; P = 0.042). CONCLUSIONS: Our study indicated that dietary folate intake had little or no effect on prostate cancer risk. However, increased serum folate levels have potentially harmful effects on the risk of prostate cancer.

Ablation of Shank1 Protects against 6-OHDA-induced Cytotoxicity via PRDX3-mediated Inhibition of ER Stress in SN4741 Cells.

BACKGROUND: Postsynaptic density (PSD) is an electron-dense structure that contains various scaffolding and signaling proteins. Shank1 is a master regulator of the synaptic scaffold located at glutamatergic synapses, and has been proposed to be involved in multiple neurological disorders. METHODS: In this study, we investigated the role of shank1 in an in vitro Parkinson's disease (PD) model mimicked by 6-OHDA treatment in neuronal SN4741 cells. The expression of related molecules was detected by western blot and immunostaining. RESULTS: We found that 6-OHDA significantly increased the mRNA and protein levels of shank1 in SN4741 cells, but the subcellular distribution was not altered. Knockdown of shank1 via small interfering RNA (siRNA) protected against 6-OHDA treatment, as evidenced by reduced lactate dehydrogenase (LDH) release and decreased apoptosis. The results of RT-PCR and western blot showed that knockdown of shank1 markedly inhibited the activation of endoplasmic reticulum (ER) stress associated factors after 6-OHDA exposure. In addition, the downregulation of shank1 obviously increased the expression of PRDX3, which was accompanied by the preservation of mitochondrial function. Mechanically, downregulation of PRDX3 via siRNA partially prevented the shank1 knockdowninduced protection against 6-OHDA in SN4741 cells. CONCLUSION: In summary, the present study has provided the first evidence that the knockdown of shank1 protects against 6-OHDA-induced ER stress and mitochondrial dysfunction through activating the PRDX3 pathway.

Qingfei mixture mitigates immunosuppression of tumor microenvironment in non-small cell lung cancer by blocking stat1/Ido1-mediated tryptophan-kynurenine pathway.

Programmed death-1 (PD-1) acts as a T cell checkpoint and is important in controlling T cell exhaustion. Blocking the intercommunication across PD-1 and PD-L1 is promising for advanced lung cancer treatment. However, the response rate requires being strengthened. This study aimed to determine whether the combination treatment of Qingfei mixture (QFM) and PD-1 inhibitor could improve the sensitivity of monoclonal antibody by regulating STAT1/IDO1-mediated tryptophan (Trp)-kynurenine (Kyn) pathway. The in vivo imaging system, immunofluorescence, hematoxylin-eosin staining, TUNEL, flow cytometry, HPLC, and ELISA were used to estimate the anti-tumor effects in LLC-luc tumor-bearing C57BL/6 mice treated with QFM, PD-1 inhibitor, 2-NP (enhancer of STAT1 transcription), and FICZ (AhR agonist) alone or in combination. IFN-γ-mediated A549 and LLC cells were treated with QFM-containing serum and fludarabine (FLU, STAT1 inhibitor), and cell viability, apoptosis, and Kyn content were then evaluated using CCK-8 assays, flow cytometry, and HPLC assays, respectively. Additionally, the expressions of STAT1, IDO1, AhR, NFATc1, TRIP12, PD-1, and PD-L1 were measured in vivo and in vitro. We found QFM increased the anti-cancer actions of PD-1 inhibitors by increasing the CD8+IFNγ+ T cells infiltration and decreasing the ratio of Kyn/Trp. Besides, QFM-containing serum suppressed the proliferation and promoted apoptosis in A549 and LLC cells, meanwhile, FLU boosted the effects of QFM-containing serum. Moreover, the suppression of tumor growth in the combination therapy was attenuated in the mice receiving 2-NP or FICZ. The occurrence of the above results was accompanied by a decrease in STAT1, IDO1, AhR, PD-1, and PD-L1 expressions. Collectively, the findings suggested that QFM may increase the influences of PD-1 inhibitors at least partially by blocking the STAT1/IDO1-mediated tryptophan-kynurenine pathway in lung cancer.

Qingfei mixture modulates the immune responses in lung cancer through modulating mTOR signaling and gut microbiota-derived short-chain fatty acids.

Lung cancer ranks among the primary contributors to cancer-related fatalities on a global scale. Multiple research investigations have demonstrated that there exists a dysbiosis within the intestinal bacteria and short-chain fatty acids (SCFAs) is linked with immune responses in lung cancer. Qingfei mixture (QFM) has been widely used in treating lung cancer, yet the active ingredients and roles of the QFM on immune responses by targeting gut microbiota remain to be elucidated. The chemical constituents of QFM were qualitatively examined by UPLC/Q-TOF-MS. Additionally, we evaluated the therapeutic impact of the organic substance QFM on lung cancer, aiming to elucidate its mechanisms for improving the tumor-immune microenvironment. Herein, we constructed a Lewis lung carcinoma (LLC)-bearing mice model with QFM treatment to observe tumor growth and immune cell changes. Then, the feces were collected and a combinatory study using metagenomes, non-targeted metabonomics, and targeted metabonomics of SCFAs was performed. In vitro experiments have been conducted to estimate the roles of acetate and sodium propionate in CD8+ T cells. Furthermore, we treated tumor-bearing mice with QFM, QFM + MHY1485 (an mTOR activator), and QFM + an antibiotic mixture (ABX) to explore the potential therapeutic benefit of regulation of the tumor microenvironment. A total of 96 compounds were obtained from QFM by UPLC/Q-TOF-MS. Besides, the findings demonstrated that QFM exhibited significant efficacy against lung cancer, manifesting in reduced tumor growth and improved immune responses. In investigating its mechanisms, we integrated gut microbiota sequencing and fecal metabolomics, revealing that QFM effectively restored disruptions in gut microbiota and SCFAs in mice with lung cancer. QFM, acetate, or sodium propionate contributed to the up-regulation of IFN-γ, Gzms-B, perforin, IL-17, IL-6, IL-12, TNF-α expressions and decreased HDAC and IL-10 levels in vitro and in vivo. Moreover, MHY1485 and ABX weakened the effects of QFM on immunomodulation. Collectively, these results suggest that QFM may facilitate immune responses in the LLC-bearing mice via regulating the gut microbiota-derived SCFAs at least partially through targeting the mTOR signaling pathway.

Transcutaneous Electrical Acupoint Stimulation Combined with Warm Acupuncture for Breast Cancer Related Upper Limb Lymphedema: A Retrospective Cohort Study.

OBJECTIVE: To observe the clinical efficacy of transcutaneous electrical acupoint stimulation (TEAS) combined with warm acupuncture in treating breast cancer associated with upper limb lymphedema (BCRL). METHODS: This was a retrospective cohort study using a paired control design. Fifty-two BCRL patients were assigned to the control group (27 cases) and the treatment group (25 cases). The patients in the control group were treated with lymphedema comprehensive detumescence treatment (CDT) for 4 weeks, including systematic therapy composed of manual lymphatic drainage, compression bandage, skincare, and functional exercise. The patients in the treatment group were treated with TEAS combined with warm acupuncture based on the control group methods. Each treatment lasted for 30 min and was applied twice a week for 4 weeks. The arm circumference (AC) of different positions of the affected limb and the degree of swelling of the affected limb were evaluated before the first treatment and after the last treatment. The clinical efficacy was evaluated according to the degree of edema before and after treatment. All adverse events during treatment were recorded. RESULTS: The patients' AC and the swelling feeling of the affected limb in the treatment group and the control group were both reduced compared with those before treatment. Compared with the control group, AC of the wrist joint transverse stria, the midpoint between the wrist joint transverse stria and the elbow joint transverse stria in the treatment group were significantly reduced (P<0.05). The decrease in AC diameter at the midpoint between the elbow joint transverse stria and the axillary transverse stria was the most significant (P<0.01). The swelling degree of the affected limbs in the treatment group was significantly lower than before treatment, and was significantly lower compared with the control group after treatment (P<0.01). The total effective rate was 72% in the treatment group, significantly higher than that in the control group (55.56%, P<0.05). No serious adverse events occured in either group. CONCLUSIONS: TEAS combined with warm acupuncture can effectively reduce AC and swelling feeling of the affected limb in patients with BCRL. The effect is better than that of CDT therapy alone. (Registration No. ChiCTR2200062075).

Qingyihuaji Formula promotes apoptosis and autophagy through inhibition of MAPK/ERK and PI3K/Akt/mTOR signaling pathway on pancreatic cancer in vivo and in vitro.

ETHNOPHARMACOLOGICAL RELEVANCE: Qingyihuaji Formula (QYHJ), a widely used traditional Chinese medicine (TCM), has been used to treat patients with cancer in China. However, the effect and mechanism of QYHJ on pancreatic ductal adenocarcinoma (PDAC) remains unclear. AIM OF THE STUDY: This study aimed to explore the roles and evaluate the possible underlying molecular mechanisms of QYHJ and its core component in PDAC using label-free quantitative proteomics in conjunction with network pharmacology-based analysis. MATERIALS AND METHODS: By screening differentially expressed proteins (DEPs) in proteomics and QYHJ-predicted gene sets, we identified QYHJ-related PDAC targets annotated with bioinformatic analysis. A subcutaneous tumor model was established to assess the role of QYHJ in vivo. The effects of quercetin (Que), a core component of QYHJ, on cell proliferation, migration, invasion, apoptosis, and autophagy in SW1990 and PANC-1 cells were investigated in vitro. Immunohistochemistry, western blotting, mRFP-GFP-LC3 adenovirus, and kinase analysis were used to determine the underlying mechanisms. RESULTS: Bioinformatics analysis revealed that 41 QYHJ-related PDAC targets were closely related to the cellular response to nitrogen compounds, positive regulation of cell death, regulation of epithelial cell apoptotic processes, and chemokine signaling pathways. CASP3, SRC, STAT1, PTPN11, PKM, and PAK1 with high expression were identified as hub DEPs in the PPI network, and these DEPs were associated with poor overall survival and STAT 1, MAPK/ERK, and PI3K/Akt/mTOR signaling pathways in PDAC patients. QYHJ significantly promoted tumor death in nude mice. Moreover, quercetin inhibited the proliferation, migration, and invasion of PDAC cells. Additionally, Que induced apoptosis and autophagy in PDAC cells. Mechanistically, QYHJ and Que significantly activated STAT 1 and remarkably inhibited the MAPK/ERK and PI3K/Akt/mTOR signaling pathways in vivo and in vitro, respectively. Importantly, ERK1/2 inactivation contributes to que-induced apoptosis in SW1990 and PANC-1 cells. CONCLUSIONS: These results suggest that QYHJ and Que are promising anti-PDAC avenues that benefit from their multiform mechanisms.

Integrated microbiome, metabolome, and proteome analysis identifies a novel interplay among commensal bacteria, metabolites and candidate targets in non-small cell lung cancer.

BACKGROUND: Accumulation of evidence suggests that the gut microbiome, its specific metabolites, and differentially expressed proteins (DEPs) are related to non-small cell lung cancer (NSCLC) pathogenesis. We now report the influences of the gut microbiota, metabolites, and DEPs on the mediation of NSCLC's chronic inflammation and immune dysregulation. METHODS: We conducted 16S ribosomal RNA sequencing for the gut microbiome in healthy volunteers and NSCLC patients. Liquid chromatography-mass spectrometry (LC-MS) analysis was employed to explore differences between metabolites and DEPs in serum samples. Additionally, LC-MS-based metabolomic analysis was conducted in 40 NSCLC tissues and 40 adjacent tissues. The omics data were separately analysed and integrated by using Spearman's correlation coefficient. Then, faecal microbiota transplantation (FMT) assay was used to assess the effects of the gut microbiome and specific metabolites in mice. RESULTS: Faecal microbiome analysis revealed gut microflora dysbiosis in NSCLC patients with Prevotella, Gemmiger, and Roseburia significantly upregulated at the genus level. Then, we identified that nervonic acid/all-trans-retinoic acid level was negatively related to Prevotella. Additionally, a total of core 8 DEPs were selected in the proteome analysis, which mainly participated in the production of IL-8 and NF-κB pathways. CRP, LBP, and CD14 were identified as potential biomarkers for NSCLC. Transplantation of faecal microbiota from patients with NSCLC or Prevotella copri-colonized recipient in mice resulted in inflammation and immune dysregulation. In turn, nervonic acid/all-trans-retinoic acid treatment improved the phenotype of C57BL/6 mice bearing P. copri-treated Lewis lung cancer (LLC). CONCLUSIONS: Overall, these results pointed out that P. copri-nervonic acid/all-trans-retinoic acid axis may contribute to the pathogenesis of NSCLC.

Application of Multigroup Technology in Non-Small-Cell Lung Cancer with Qi Stagnation and Blood Stasis Syndrome.

OBJECTIVE: To explore the basic characteristics of intestinal flora, metabolomics, and proteomics of non-small cell lung cancer (NSCLC) in patients with Qi stagnation and blood stasis syndrome. METHODS: Twelve NSCLC patients with Qi stagnation and blood stasis syndrome were selected for the QZXY group and 15 healthy volunteers were selected for the control group. Fecal samples from the two groups were collected to evaluate intestinal microecology using the 16s rDNA technique. Serum samples were collected to compare the differences in metabolomics and proteomics between the two groups using liquid chromatography-mass spectrometry (LC-MS). Another 34 NSCLC patients with other syndromes were selected for the nQZXY group and their serum samples were collected. Metabolomics differences between the QZXY and nQZXY groups were compared using LC-MS, and four metabolites with the most obvious differences were selected for receiver operation characteristic curve representation. Finally, multigroup results were analyzed using the WGCNA software. RESULTS: There were two significantly different types of bacteria (Aerococcaceae and Abiotrophia), 11 different proteins (six upregulated and five downregulated), and 38 different metabolites (nine upregulated, 29 downregulated) between the QZXY and control groups. There was a correlation between differential bacteria, proteins, and metabolites. The conjoint analysis found that the different substances were related to MAPK, PI3K/Akt, Ras signaling pathway, cancer pathways, and cytokine-cytokine receptor interaction. There were four significant differences in metabolites (Pseudouridine, phenlacetyl-C0A, L-glutamic, and phospho-anandamide) between the QZXY and nQZXY groups. CONCLUSIONS: NSCLC with Qi stagnation and blood stasis syndrome had specific intestinal flora and protein and metabolites, which were closely related to the occurrence and development of tumors.