Analysis of intracellular communication reveals consistent gene changes associated with early-stage acne skin.

A comprehensive understanding of the intricate cellular and molecular changes governing the complex interactions between cells within acne lesions is currently lacking. Herein, we analyzed early papules from six subjects with active acne vulgaris, utilizing single-cell and high-resolution spatial RNA sequencing. We observed significant changes in signaling pathways across seven different cell types when comparing lesional skin samples (LSS) to healthy skin samples (HSS). Using CellChat, we constructed an atlas of signaling pathways for the HSS, identifying key signal distributions and cell-specific genes within individual clusters. Further, our comparative analysis revealed changes in 49 signaling pathways across all cell clusters in the LSS- 4 exhibited decreased activity, whereas 45 were upregulated, suggesting that acne significantly alters cellular dynamics. We identified ten molecules, including GRN, IL-13RA1 and SDC1 that were consistently altered in all donors. Subsequently, we focused on the function of GRN and IL-13RA1 in TREM2 macrophages and keratinocytes as these cells participate in inflammation and hyperkeratinization in the early stages of acne development. We evaluated their function in TREM2 macrophages and the HaCaT cell line. We found that GRN increased the expression of proinflammatory cytokines and chemokines, including IL-18, CCL5, and CXCL2 in TREM2 macrophages. Additionally, the activation of IL-13RA1 by IL-13 in HaCaT cells promoted the dysregulation of genes associated with hyperkeratinization, including KRT17, KRT16, and FLG. These findings suggest that modulating the GRN-SORT1 and IL-13-IL-13RA1 signaling pathways could be a promising approach for developing new acne treatments.

Gasdermin D Inhibitor Necrosulfonamide Alleviates Angiotensin II-Induced Abdominal Aortic Aneurysms in Apolipoprotein E-Deficient Mice.

Abdominal aortic aneurysm (AAA) is a chronic aortic disease that lacks effective pharmacological therapies. This study was performed to determine the influence of treatment with the gasdermin D inhibitor necrosulfonamide on experimental AAAs. AAAs were induced in male apolipoprotein E-deficient mice by subcutaneous angiotensin II infusion (1000 ng/kg body weight/min), with daily administration of necrosulfonamide (5 mg/kg body weight) or vehicle starting 3 days prior to angiotensin II infusion for 30 days. Necrosulfonamide treatment remarkably suppressed AAA enlargement, as indicated by reduced suprarenal maximal external diameter and surface area, and lowered the incidence and reduced the severity of experimental AAAs. Histologically, necrosulfonamide treatment attenuated medial elastin breaks, smooth muscle cell depletion, and aortic wall collagen deposition. Macrophages, CD4+ T cells, CD8+ T cells, and neovessels were reduced in the aneurysmal aortas of necrosulfonamide- as compared to vehicle-treated angiotensin II-infused mice. Atherosclerosis and intimal macrophages were also substantially reduced in suprarenal aortas from angiotensin II-infused mice following necrosulfonamide treatment. Additionally, the levels of serum interleukin-1ß and interleukin-18 were significantly lower in necrosulfonamide- than in vehicle-treated mice without affecting body weight gain, lipid levels, or blood pressure. Our findings indicate that necrosulfonamide reduced experimental AAAs by preserving aortic structural integrity as well as reducing mural leukocyte accumulation, neovessel formation, and systemic levels of interleukin-1ß and interleukin-18. Thus, pharmacologically inhibiting gasdermin D activity may lead to the establishment of nonsurgical therapies for clinical AAA disease.

Investigating cellular similarities and differences between upper tract urothelial carcinoma and bladder urothelial carcinoma using single-cell sequencing.

Background: Upper tract urothelial carcinoma (UTUC) and bladder urothelial carcinoma (BLCA) both originate from uroepithelial tissue, sharing remarkably similar clinical manifestations and therapeutic modalities. However, emerging evidence suggests that identical treatment regimens may lead to less favorable outcomes in UTUC compared to BLCA. Therefore, it is imperative to explore molecular processes of UTUC and identify biological differences between UTUC and BLCA. Methods: In this study, we performed a comprehensive analysis using single-cell RNA sequencing (scRNA-seq) on three UTUC cases and four normal ureteral tissues. These data were combined with publicly available datasets from previous BLCA studies and RNA sequencing (RNA-seq) data for both cancer types. This pooled analysis allowed us to delineate the transcriptional differences among distinct cell subsets within the microenvironment, thus identifying critical factors contributing to UTUC progression and phenotypic differences between UTUC and BLCA. Results: scRNA-seq analysis revealed seemingly similar but transcriptionally distinct cellular identities within the UTUC and BLCA ecosystems. Notably, we observed striking differences in acquired immunological landscapes and varied cellular functional phenotypes between these two cancers. In addition, we uncovered the immunomodulatory functions of vein endothelial cells (ECs) in UTUC, and intercellular network analysis demonstrated that fibroblasts play important roles in the microenvironment. Further intersection analysis showed that MARCKS promote UTUC progression, and immunohistochemistry (IHC) staining revealed that the diverse expression patterns of MARCKS in UTUC, BLCA and normal ureter tissues. Conclusion: This study expands our multidimensional understanding of the similarities and distinctions between UTUC and BLCA. Our findings lay the foundation for further investigations to develop diagnostic and therapeutic targets for UTUC.

A regressive analysis of the main environmental risk factors of human echinococcosis in 370 counties in China.

BACKGROUND: Echinococcosis is a natural focal, highly prevalent disease in China. Factors influencing the spread of echinococcosis are not only related to personal exposure but also closely related to the environment itself. The purpose of this study was to explore the influence of environmental factors on the prevalence of human echinococcosis and to provide a reference for prevention and control of echinococcosis in the future. METHODS: Data were collected from 370 endemic counties in China in 2018. By downloading Modis, DEM and other remote-sensing images in 2018. Data on environmental factors, i.e., elevation, land surface temperature (LST) and normalized difference vegetation index (NDVI) were collected. Rank correlation analysis was conducted between each environmental factor and the prevalence of echinococcosis at the county level. Negative binomial regression was used to analyze the impact of environmental factors on the prevalence of human echinococcosis at the county level. RESULTS: According to rank correlation analysis, the prevalence of human echinococcosis in each county was positively correlated with elevation, negatively correlated with LST, and negatively correlated with NDVI in May, June and July. Negative binomial regression showed that the prevalence of human echinococcosis was negatively correlated with annual LST and summer NDVI, and positively correlated with average elevation and dog infection rate. The prevalence of human cystic echinococcosis was inversely correlated with the annual average LST, and positively correlated with both the average elevation and the prevalence rate of domestic animals. The prevalence of human alveolar echinococcosis was positively correlated with both NDVI in autumn and average elevation, and negatively correlated with NDVI in winter. CONCLUSION: The prevalence of echinococcosis in the population is affected by environmental factors. Environmental risk assessment and prediction can be conducted in order to rationally allocate health resources and improve both prevention and control efficiency of echinococcosis.

Comparison of EUS-FNA and EUS-FNB for diagnosis of solid pancreatic mass lesions: a meta-analysis of prospective studies.

Objective: To quantitatively compare the diagnostic value of endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) and endoscopic ultrasound-guided fine needle biopsy (EUS-FNB) in solid pancreatic mass lesions using a systematic evaluation method.Methods: A systematic literature search was conducted on public databases to include studies comparing the diagnostic value of EUS-FNA and EUS-FNB in solid pancreatic mass lesions. The combined effect size was estimated using mean difference (MD) and risk difference (RD) respectively, and the corresponding 95% confidence interval (CI) was calculated.Results: The 12 articles (7 RCTs and 5 cohort studies) met the inclusion criteria of this study. The meta-analysis showed that compared with EUS-FNB, EUS-FNA had lower diagnostic accuracy (RD: -0.08, 95% CI: -0.15, -0.01) and specimen adequacy (RD: -0.08, 95% CI: -0.15, -0.02), while higher required number of needle passes (MD: 0.42, 95% CI: 0.12, 0.73). However, EUS-FNB and EUS-FNA presented similar overall complications (RD: 0.00, 95% CI: -0.01, 0.02) and technical failures (RD: -0.01, 95% CI: -0.02, 0.00), without statistically significant differences.Conclusions: Compared with EUS-FNA, EUS-FNB seems to be a better choice for diagnosing suspected pancreatic lesions.

Biocontrol and Growth Promotion Potential of Bacillus subtilis CTXW 7-6-2 against Rhizoctonia solani that Causes Tobacco Target Spot Disease.

Fungal diseases form perforated disease spots in tobacco plants, resulting in a decline in tobacco yield and quality. The present study investigated the antagonistic effect of Bacillus subtilis CTXW 7-6-2 against Rhizoctonia solani, its ability to promote the growth of tobacco seedlings, and the expression of disease resistance-related genes for efficient and eco-friendly plant disease control. Our results showed that CTXW 7-6-2 had the most vigorous growth after being cultured for 96 h, and its rate of inhibition of R. solani growth in vitro was 94.02%. The volatile compounds produced by CTXW 7-6-2 inhibited the growth of R. solani significantly (by 96.62%). The fungal growthinhibition rate of the B. subtilis CTXW 7-6-2 broth obtained after high-temperature and no-high-temperature sterile fermentation was low, at 50.88% and 54.63%, respectively. The lipopeptides extracted from the B. subtilis CTXW 7-6-2 fermentation broth showed a 74.88% fungal growth inhibition rate at a concentration of 100 mg/l. Scanning and transmission electron microscopy showed some organelle structural abnormalities, collapse, shrinkage, blurring, and dissolution in the R. solani mycelia. In addition, CTXW 7-6-2 increased tobacco seedling growth and improved leaf and root weight compared to the control. After CTXW 7-6-2 inoculation, tobacco leaves showed the upregulation of the PDF1.2, PPO, and PAL genes, which are closely related to target spot disease resistance. In conclusion, B. subtilis CTXW 7-6-2 may be an efficient biological control agent in tobacco agriculture and enhance plant growth potential.

LATS2 condensates organize signalosomes for Hippo pathway signal transduction.

Biomolecular condensates have been proposed to mediate cellular signaling transduction. However, the mechanism and functional consequences of signal condensates are not well understood. Here we report that LATS2, the core kinase of the Hippo pathway, responds to F-actin cytoskeleton reduction and forms condensates. The proline-rich motif (PRM) of LATS2 mediates its condensation. LATS2 partitions with the main components of the Hippo pathway to assemble a signalosome for LATS2 activation and for its stability by physically compartmentalizing from E3 ligase FBXL16 complex-dependent degradation, which in turn mediates yes-associated protein (YAP)-transcriptional coactivator with PDZ-binding motif (TAZ) recruitment and inactivation. This oncogenic FBXL16 complex blocks LATS2 condensation by binding to the PRM region to promote its degradation. Disruption of LATS2 condensation leads to tumor progression. Thus, our study uncovers that the signalosomes assembled by LATS2 condensation provide a compartmentalized and reversible platform for Hippo signaling transduction and protein stability, which have potential implications in cancer diagnosis and therapeutics.

Immunotoxicity Evaluation of Trihalophenolic Disinfection By-Products in Mouse and Human Mononuclear Macrophage Systems: The Role of RNA Epitranscriptomic Modification in Mammalian Immunity.

BACKGROUND: 2,4,6-Trichlorophenol (TCP), 2,4,6-tribromophenol (TBP) and 2,4,6-triiodophenol (TIP) are three widely detected trihalophenolic disinfection by-products (DBPs). Previous studies have mainly focused on the carcinogenic risk and developmental toxicity of 2,4,6-trihalophenols. Very little is known about their immunotoxicity in mammals. OBJECTIVES: We investigated the effects of 2,4,6-trihalophenols on mammalian immunity using a mouse macrophage model infected with bacteria or intracellular parasites and aimed to elucidate the underlying mechanisms from an epitranscriptomic perspective. The identified mechanisms were further validated in human peripheral blood mononuclear cells (PBMCs). METHODS: The mouse macrophage cell line RAW264.7 and primary mouse peritoneal macrophages were exposed to different concentrations of TCP, TBP, and TIP. The pro-inflammatory marker Ly6C, the survival of the bacterium Escherichia coli (E. coli), and the parasite burden of Toxoplasma gondii (T. gondii) were assessed. Furthermore, the global gene expression profiling of macrophages following exposure to 2,4,6-trihalophenols was obtained through RNA-sequencing (RNA-seq). The effects of 2,4,6-trihalophenols on RNA N6-methyladenosine (m6A) methyltransferases and total RNA m6A levels were evaluated using Western blotting and dot blot, respectively. Transcriptome-wide m6A methylome was analyzed by m6A-seq. In addition, expression of m6A regulators and total RNA m6A levels in human PBMCs exposed to 2,4,6-trihalophenols were detected using quantitative reverse transcriptase polymerase chain reaction and dot blot, respectively. RESULTS: Mouse macrophages exposed to TCP, TBP, or TIP had lower expression of the pro-inflammatory marker Ly6C, with a greater difference from control observed for TIP-exposed cells. Consistently, macrophages exposed to such DBPs, especially TIP, were susceptible to infection with the bacterium E. coli and the intracellular parasite T. gondii, indicating a compromised ability of macrophages to defend against pathogens. Intriguingly, macrophages exposed to TIP had significantly greater m6A levels, which correlated with the greater expression levels of m6A methyltransferases. Macrophages exposed to each of the three 2,4,6-trihalophenols exhibited transcriptome-wide redistribution of m6A. In particular, the m6A peaks in genes associated with immune-related pathways were altered after exposure. In addition, differences in m6A were also observed in human PBMCs after exposure to 2,4,6-trihalophenols. DISCUSSION: These findings suggest that 2,4,6-trihalophenol exposure impaired the ability of macrophages to defend against pathogens. This response might be associated with notable differences in m6A after exposure. To the best of our knowledge, this study presents the first m6A landscape across the transcriptome of immune cells exposed to pollutants. However, significant challenges remain in elucidating the mechanisms by which m6A mediates immune dysregulation in infected macrophages after 2,4,6-trihalophenol exposure. https://doi.org/10.1289/EHP11329.

Pharmacological Inhibition of Gasdermin D Suppresses Angiotensin II-Induced Experimental Abdominal Aortic Aneurysms.

BACKGROUND: Gasdermin D, a molecule downstream of the nucleotide-binding oligomerization domain-like receptor family pyrin domain containing inflammasome, forms the membrane pore for the secretion of interleukin (IL)-1ß and IL-18, and also mediates pyroptosis. This study was to explore the influence of treatment with disulfiram, a small molecule inhibitor to gasdermin D, on the formation and progression of experimental abdominal aortic aneurysms (AAA). METHODS: AAAs were induced in 10-week-old male apolipoprotein E deficient mice by subcutaneous infusion of angiotensin II (1000 ng/min/kg body weight) for 28 days via osmotic minipumps. Three days prior to angiotensin II infusion, disulfiram (50 mg/kg) or an equal volume of saline as the vehicle control was administered daily via oral gavage. The influence on experimental AAAs was analyzed by serial measurements of aortic diameters via ultrasonography, grading AAA severity and histopathology at sacrifice. Serum IL-1ß and IL-18 levels, systolic blood pressure, total cholesterol, and triglyceride were also measured. Additional experiments assayed the influences on the cell viability and IL-1ß secretion of in vitro activated macrophages. RESULTS: Disulfiram significantly reduced the enlargement, incidence, and severity of angiotensin II-induced experimental AAAs with attenuation of medial elastin breaks, mural macrophage accumulation, and systolic blood pressure. The AAA suppression was also associated with reduced systemic levels of IL-1ß but not IL-18. However, disulfiram treatment had no impact on body weight gain and lipid levels in aneurysmal mice. Additionally, disulfiram treatment also markedly reduced the secretion of IL-1ß from activated macrophages with a limited effect on cell viability in vitro. CONCLUSIONS: Gasdermin D inhibition by disulfiram attenuated angiotensin II-induced experimental AAAs with reduced systemic IL-1ß levels and in vitro activated macrophage IL-1ß secretion. Our study suggests that pharmacological gasdermin D inhibition may have translational potential for limiting clinical AAA progression.

Toxoplasma gondii microneme protein MIC3 induces macrophage TNF-α production and Ly6C expression via TLR11/MyD88 pathway.

Toxoplasma gondii is the most successful parasite worldwide. It is of great interest to understand how T. gondii induce different immune responses in different hosts. In this study, we found that a peptide of T. gondii microneme protein MIC3 induced TNF-α production, NF-κB phosphorylation, iNOS transcription and Ly6C expression in mouse macrophage RAW264.7 cells. MyD88 inhibition, small interfering RNA against Tlr11 and CRISPR/Cas9-mediated knock-out of Tlr11 all reduced MIC3-induced TNF-α production, NF-κB phosphorylation, iNOS transcription and Ly6C expression. Additionally, we determined the location of MIC3 peptide in mouse macrophages using immunofluorescence. MIC3 could both adhere to the cell membrane of mouse macrophages and enter the cells. These results suggest that MIC3 triggered the immune responses in mouse macrophages via TLR11/MyD88/NF-κB pathway. It is known that human macrophages lacking TLR11. We predicted that the immune responses induced by MIC3 in human macrophages were significantly different from those in mouse macrophages. As expected, MIC3 peptide failed to induce TNF-α expression, iNOS expression and NF-κB phosphorylation in human THP-1 derived macrophages. MIC3 induced macrophage immune responses via TLR11. Intriguingly, the amino acid sequence of MIC3 is completely different from the well-known TLR11 ligand profilin, which generates a potent IL-12p40, TNF-α and IL-6 response. In marked contrast to profilin, MIC3 could not induce IL-12p40 expression in both mouse RAW264.7 cells and human THP-1 derived macrophages. Furthermore, the simulated tertiary structure of MIC3 peptide shows poor similarity with the crystal structure of profilin, suggesting that MIC3 might be a different ligand from profilin. These findings about MIC3 and TLR11 will provide us with important insights into the pathogenesis of toxoplasmosis and coevolution during host-parasite interaction.

Investigation of the Active Compounds and Important Pathways of Huaiqihuang Granule for the Treatment of Immune Thrombocytopenia Using Network Pharmacology and Molecular Docking.

Materials and Methods: Compounds of HQHG were scanned by LC-MS/MS, and the target profiles of compounds were identified based on SwissTarget Prediction. ITP target proteins were collected from various databases. Then, KEGG pathway and GO enrichment analyses were performed to explore the signaling pathways related to HQHG for ITP. The PPI and drug-herbs-compounds-targets-pathways network were constructed using Cytoscape 3.7.2. Finally, Discovery studio software was used to confirm the key targets and active compounds from HQHG. Results: A total of 187 interacting targets of 19 potentially active compounds in HQHG and 3837 ITP-related targets were collected. Then, 187 intersection targets were obtained. A total of 20 key targets including EGFR, CASP3, TNF, STAT3, and ERBB2 were identified through PPI network analysis. These targets were mainly focused on the biological processes of positive regulation of protein phosphorylation, cellular response to organonitrogen compound, and cellular response to nitrogen compound. 20 possible pathways of HQHG in the treatment of ITP were identified through KEGG enrichment. EGFR, CASP3, TNF, and STAT3 are the four most important target proteins, while adenosine, caffeic acid, ferulic acid, quercetin-3ß-D-glucoside, rutin, scopoletin, and tianshic acid are the most important active compounds, which were validated by molecular docking simulation. Conclusion: This study demonstrated that HQHG produced relief effects against ITP by regulating multitargets and multipathways with multicompounds. And the combined data provide novel insight of drug developing for ITP.

Assessing OBT formation and enrichment: ROS signaling is involved in the radiation hormesis induced by tritium exposure in algae.

Tritium is the main component of radioactive wastewater from nuclear power plants and can be migrated into organisms to form organically bound tritium (OBT), which may pose a potential risk to aquatic ecosystem. Hence, it is essential to monitor OBT conversion in the presence of tritium exposure. In this study, the effects of pretreatment methods such as digestion on the recovery of tritium were discussed. It was found that microwave digestion pretreatment could improve the recovery of tritium by up to 90 % and allow OBT measurement with a small sample size equivalent to about 60 mg (dry weight). In addition, the efficiency of OBT transformation was different among biological samples, and the radiation hormesis phenomenon was induced by tritium exposure (3.7 × 106 Bq/L) in microalgae Chlorella vulgaris（C. vulgaris）. The tritium exposure may induce radiation hormesis through the reactive oxygen species (ROS) signaling pathway, thus improving the photosynthetic capacity and metabolism level of C. vulgaris. Furthermore, enhancement of photorespiration metabolism and the antioxidation system may be important means for C. vulgaris to balance damage by tritium radiation. This study provides insights for further investigating OBT behavior, and will contribute to understanding the equilibrium damage mechanism of algae exposed to tritium.

Societal drivers of human echinococcosis in China.

BACKGROUND: Echinococcosis is a parasitic zoonotic disease that threatens human health and economic development. In China, 370 counties are endemic for echinococcosis. Qinghai-Tibet Plateau has the most patients and people at risk. Therefore, analyzing the societal factors related to susceptibility to the disease is critical for efficient prevention and control of echinococcosis. METHODS: The demographic characteristics and lifestyle of echinococcosis cases were clustered using K-means cluster analysis to determine the main factors of risk of echinococcosis. RESULTS: Middle-aged and young people as well as those with a low education level and herdsmen are at risk of contracting echinococcosis. Nomadism, domestic and feral dogs in the surrounding environment, and drinking heavily polluted natural surface water are the main behavioral risk factors. The cystic echinococcosis (CE) and alveolar echinococcosis (AE) cluster analysis focused on female, middle-aged, and young people, winter settlement and summer nomadism, and domestic and feral dogs in the surrounding environment. There were significant differences in lifestyle between Qinghai-Tibet Plateau cases and non-Qinghai-Tibet-Plateau cases. CONCLUSION: According to the distribution of cases and CE and AE, this study identified the factors of risk of echinococcosis in the Qinghai-Tibet Plateau and non-Qinghai-Tibet Plateau. Adapted control techniques appropriate for the various epidemic areas should be established to serve as a reference for echinococcosis prevention.

Global regulatory factor AaLaeA upregulates the production of antitumor substances in the endophytic fungus Alternaria alstroemeria.

The global regulatory factor LaeA has been shown to be involved in the biosynthesis of secondary metabolites in various fungi. In a previous work, we isolated an endophytic fungus from Artemisia annua, and its extract had a significant inhibitory effect on the A549 cancer cell line. Phylogenetic analysis further identified the strain as Alternaria alstroemeria. Overexpression of AalaeA gene resulted in significantly increased antitumor activity of this strain's extract. The 3-(4, 5- dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay results showed that the inhibition rate of the AalaeAOE29 mutant extract on A549 cancer cells was significantly higher than that of the WT extract, as the IC50 decreased from 195.0 to 107.4 µg/ml, and the total apoptosis rate was enhanced. Overexpression of the AalaeA gene significantly increased the contents of myricetin, geraniol, ergosterol, and 18 other antitumor compounds as determined by metabolomic analysis. Transcriptomic analysis revealed significant changes in 95 genes in the mutant strain, including polyketide synthases, nonribosomal peptide synthases, cytochrome P450s, glycosyltransferases, acetyl-CoA acetyltransferases, and others. These results suggested that AaLaeA mediated the antitumor activity of the metabolites in A. alstroemeria by regulating multiple metabolic pathways.

Connexin32 regulates expansion of liver cancer stem cells via the PI3K/Akt signaling pathway.

Liver cancer stem cells (LCSCs) are responsible for liver cancer recurrence, metastasis, and drug resistance. Previous studies by the authors demonstrated that upregulated expression of connexin 32 (Cx32) reversed doxorubicin resistance and reduced invasion and metastasis of liver cancer cells. However, the role of Cx32 in expansion of LCSCs remains unclear. A total of 85 patients were enrolled in the present study and followed­up for 5 years. The expression of Cx32 in hepatocellular carcinoma (HCC) tissues and corresponding paracancerous tissues were detected by immunohistochemistry (IHC). Cx32 was silenced in HepG2 cells and overexpressed in HCCLM3 cells and the stemness of liver cells was examined by detecting the expression of LCSC markers (EpCAM, CD133, Nanog, Oct4, Sox9, c­Myc), sphere formation, and xenograft tumorigenesis. Finally, the effect of the phosphoinositide 3­kinase (PI3K)/protein kinase B (Akt) pathway on Cx32­regulated LCSC expansion was investigated. Cx32 was downregulated in LCSCs and HCC tissues, and predicted poor prognosis in patients with HCC. Overexpression of Cx32 in HCCLM3 cells significantly inhibited LCSC expansion, tumorigenesis, and phosphoinositide 3­kinase/protein kinase B (PI3K/Akt) pathway activity. By contrast, silencing of Cx32 in HepG2 cells upregulated expansion of LCSCs and PI3K/Akt pathway activity. Modulating the activity of the PI3K/Akt pathway by SC­79 and LY294002 in HepG2 and HCCLM3 cells, respectively, confirmed that Cx32 could affect the expansion of LCSCs through PI3K/Akt signaling. In conclusion, the present study demonstrated that Cx32 regulated the expansion of LCSCs, and increased expression of Cx32 significantly inhibited the expansion of LCSCs, suggesting that Cx32 may be an optimal target for intervention of HCC.

Global regulatory factor VeA upregulates the production of antitumor substances in endophytic Fusarium solani.

A number of studies have demonstrated that endophytic fungi have the potential to produce antitumor active substances with novel structures and significant activities. In our previous studies, we isolated a Fusarium strain from the stem of the medicinal plant Nothapodytes pittosporoides (Oliv.). In this study, we identified this strain as Fusarium solani and found that its crude extract has significant antitumor activity against human alveolar adenocarcinoma cells (A549). We overexpressed the global regulatory factor VeA in F. solani (VeAOE), resulting in a significant increase in antitumor activity. The MTT assay results showed that the inhibition rate of the VeAOE mutant extract on A549 cancer cells was significantly higher than that of the WT extract, as the IC50 decreased from 369.22 to 285.89 µg/mL, and the apoptosis ratio was significantly increased by approximately 4.86-fold. In VeAOE, accumulation of alkaloids, terpenoids, carboxylic acid derivatives, phenols and flavonoid metabolites with potential antitumor activity was significantly increased compared with WT based on metabolomic analysis. Additionally, transcriptome analysis found that the expression patterns of 48 genes related to antitumor activity were significantly changed in VeAOE, mainly involving glycosyl hydrolases, the Zn(2)-Cys(6) class, cytochrome P450 monooxygenase, 3-isopropylmalate dehydratase, and polyketide synthases. These results suggested that VeA mediated the antitumor activity of the metabolites in F. solani HB1-J1 by regulating multiple metabolic pathways.

Overexpression of the global regulator FnVeA upregulates antitumor substances in endophytic Fusarium nematophilum.

The special niche of endophytic fungi promotes their potential to produce antitumor compounds with novel structure and significant bioactivity for screening of new antitumor drugs. In our previous studies, we isolated a Fusarium strain from the roots of the medicinal plant Nothapodytes pittosporoides and identified it as Fusarium nematophilum. We found that the crude extract of F. nematophilum had significant antitumor activity on A549 cancer cells, and overexpressing the global regulatory factor FnVeA (the VeA gene of the fungus F. nematophilum) resulted in a significant increase in the antitumor activity, which was approximately fivefold higher than wild strain for relative inhibition rate. In FnVeAOE, the accumulation of indole, alkene, alkaloid, steroid, and flavonoid metabolites with potential antitumor activity was significantly upregulated compared with wild type via metabolomic analysis. Moreover, the transcriptome analysis showed that 134 differential genes were considered to be closely related to the biosynthesis of antitumor substances, of which 59 differential genes were considered as candidate key genes, and related to tryptophan dimethylallyltransferase, cytochrome P450 monooxygenase, polyketide synthases, and transcription factors. Taken together, we suggest that FnVeA may regulate the biosynthesis of antitumor substances by mediating the expression of genes related to secondary metabolic pathways in F. nematophilum.

Assessment of echinococcosis control in Tibet Autonomous Region, China.

BACKGROUND: In China the highest prevalence of echinococcosis is in Tibet Autonomous Region (TAR). The government has issued documents and implemented comprehensive prevention and control measures focusing on controlling the source of infection of echinococcosis. It was very important to understand the implementation and effect of infectious source control measures. The purpose of this study was to examine the implementation of measures to control infectious source (domestic and stray dogs) in TAR and to assess their effectiveness. METHODS: We collected data on domestic dog registration and deworming and stray dog sheltering in 74 counties/districts in the TAR from 2017 to 2019. Fecal samples from domestic dogs were collected from randomly selected towns to determine Echinococcus infection in dogs using coproantigen ELISA. We analyzed the data to compare the canine rate of infection between 2016 and 2019. The data analysis was performed by SPSS statistical to compare dog infection rate in 2016 and 2019 by chi-square test, and ArcGIS was used for mapping. RESULTS: From 2017 to 2019, 84 stray dog shelters were built in TAR, and accumulatively 446,660 stray or infected dogs were arrested, sheltered, or disposed of. The number of domestic dogs went downward, with an increased registration management rate of 78.4% (2017), 88.8% (2018), and 99.0% (2019). Dogs were dewormed 5 times in 2017, 12 times in 2018, and 12 times in 2019. The dog infection rate was 1.7% (252/14,584) in 2019, significantly lower than 7.3% (552/7564) from the survey of echinococcosis prevalence in Tibet in 2016 (P < 0.05). CONCLUSION: Between 2017 and 2019, the number of stray dogs and infection rate of Echinococcus spp. in domestic dogs decreased significantly, indicating that dogs were effectively controlled as a source of infection in TAR and reflecting a significant decrease in the risk of echinococcosis transmission.

Cloning and Functional Characterization of Two Germacrene A Oxidases Isolated from Xanthium sibiricum.

Sesquiterpene lactones (STLs) from the cocklebur Xanthium sibiricum exhibit significant anti-tumor activity. Although germacrene A oxidase (GAO), which catalyzes the production of Germacrene A acid (GAA) from germacrene A, an important precursor of germacrene-type STLs, has been reported, the remaining GAOs corresponding to various STLs' biosynthesis pathways remain unidentified. In this study, 68,199 unigenes were studied in a de novo transcriptome assembly of X. sibiricum fruits. By comparison with previously published GAO sequences, two candidate X. sibiricum GAO gene sequences, XsGAO1 (1467 bp) and XsGAO2 (1527 bp), were identified, cloned, and predicted to encode 488 and 508 amino acids, respectively. Their protein structure, motifs, sequence similarity, and phylogenetic position were similar to those of other GAO proteins. They were most strongly expressed in fruits, according to a quantitative real-time polymerase chain reaction (qRT-PCR), and both XsGAO proteins were localized in the mitochondria of tobacco leaf epidermal cells. The two XsGAO genes were cloned into the expression vector for eukaryotic expression in Saccharomyces cerevisiae, and the enzyme reaction products were detected by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS) methods. The results indicated that both XsGAO1 and XsGAO2 catalyzed the two-step conversion of germacrene A (GA) to GAA, meaning they are unlike classical GAO enzymes, which catalyze a three-step conversion of GA to GAA. This cloning and functional study of two GAO genes from X. sibiricum provides a useful basis for further elucidation of the STL biosynthesis pathway in X. sibiricum.

Geographic distribution and prevalence of human echinococcosis at the township level in the Tibet Autonomous Region.

BACKGROUND: Echinococcosis, a zoonotic parasitic disease, is caused by larval stages of cestodes in the Echinococcus genus. Echinococcosis is highly prevalent in ten provinces/autonomous regions of western and northern China. In 2016, an epidemiological survey of Tibet Autonomous Region (TAR) revealed that the prevalence of human echinococcosis was 1.66%, which was much higher than the average prevalence in China (0.24%). Therefore, to improve on the current prevention and control measures, it is important to understand the prevalence and spatial distribution characteristics of human echinococcosis at the township level in TAR. METHODS: Data for echinococcosis cases in 2018 were obtained from the annual report system of echinococcosis of Tibet Center for Disease Control and Prevention. Diagnosis had been performed via B-ultrasonography. The epidemic status of echinococcosis in all townships in TAR was classified according to the relevant standards of population prevalence indices as defined in the national technical plan for echinococcosis control. Spatial scan statistics were performed to establish the geographical townships that were most at risk of echinococcosis. RESULTS: In 2018, a total of 16,009 echinococcosis cases, whose prevalence was 0.53%, were recorded in 74 endemic counties in TAR. Based on the order of the epidemic degree, all the 692 townships were classified from high to low degrees. Among them, 127 townships had prevalence rates ≥ 1%. The high prevalence of human echinococcosis in TAR, which is associated with a wide geographic distribution, is a medical concern. Approximately 94.65% of the villages and towns reported echinococcosis cases. According to spatial distribution analysis, the prevalence of human echinococcosis was found to be clustered, with the specific clustering areas being identified. The cystic echinococcosis primary cluster covered 88 townships, while that of alveolar echinococcosis's covered 38 townships. CONCLUSIONS: This study shows spatial distributions of echinococcosis with different epidemic degrees in 692 townships of TAR and high-risk cluster areas at the township level. Our findings indicate that strengthening the echinococcosis prevention and control strategies in TAR should directed at townships with a high prevalence and high-risk clustering areas.