ALDH1A1 Activity in Tumor-Initiating Cells Remodels Myeloid-Derived Suppressor Cells to Promote Breast Cancer Progression.

Tumor-initiating cells (TIC) are associated with tumor initiation, growth, metastasis, and recurrence. Aldehyde dehydrogenase 1A1 (ALDH1A1) is a TIC marker in many cancers, including breast cancer. However, the molecular mechanisms underlying ALDH1A1 functions in solid tumors remain largely unknown. Here we demonstrate that ALDH1A1 enzymatic activity facilitates breast tumor growth. Mechanistically, ALDH1A1 decreased the intracellular pH in breast cancer cells to promote phosphorylation of TAK1, activate NFκB signaling, and increase the secretion of GM-CSF, which led to myeloid-derived suppressor cell expansion and immunosuppression. Furthermore, the ALDH1A1 inhibitor disulfiram and chemotherapeutic agent gemcitabine cooperatively inhibited breast tumor growth and tumorigenesis by purging ALDH+ TICs and activating T-cell immunity. These findings elucidate how active ALDH1A1 modulates the immune system to promote tumor development, highlighting new therapeutic strategies for malignant breast cancer. SIGNIFICANCE: ALDH1A1 enzyme activity induces MDSC expansion and triggers a procancer immune microenvironment to facilitate breast cancer progression, providing a novel therapeutic vulnerability in this disease.

[Enhanced epitope immunoreactivity of the dominant epitope of Toxoplasma gondii fused at the "N terminus" of HPV16L1].

For improving epitope immunogenicity and achieving the co-immunization, late protein 1 (L1) of HPV type 16 (HPV16L1) was selected as the vector to carry the dominant epitope of Toxoplasma gondii because of the shared common population between Toxoplasma gondii and human papillomavirus (HPV). RSepitope-HPV16L1 (RSepitope fused at the "N-terminus" of HPV16L1) and HPV16L1-RSepitope (RSepitope fused at the "C-terminus" of HPV16L1) chimeras were constructed. After transfection of COS-7 cells with the recombinants, Western blot, RT-PCR, and immunofluorescence experiments confirmed that RSepitope-HPV16L1 could successfully express the corresponding mRNA and protein of RSepitope and HPV16L1, but the HPV16L1-RSepitope construct could not. A "prime-boost" immunization program was applied in mice to further evaluate the immune response elicited by the constructs, and the RSepitope-HPV16L1 immunization group produced the most significantly increased humoral and cellular immune responses (the highest RSepitope-specific IgG antibody level and the highest IFN-γ production, respectively), in which both elevated Th1 and Th2 immune responses were obtained. Moreover, the advantage of HPV16L1 as an epitope carrier was remarkable for RSepitope-HPV16L1, which induced a more prominent immunological response than RSepitope alone (without fusion with HPV16L1). Our research indicated that the N-terminus of HPV16L1 could be a better insertion site for enhancing target epitope immunogenicity, and our study offers a design for epitope vaccine of reasonable combination.

Gene expression profiling of papaya (Carica papaya L.) immune response induced by CTS-N after inoculating PLDMV.

Plant immune regulation is a defensive strategy of plants for protection against pathogen invasion, and Chitosan-N (CTS-N) can induce plant autoimmunity regulation mechanisms. CTS-N was found to induce an immunomodulatory response in papaya against Papaya leaf-distortion mosaic virus (PLDMV). To date, the gene expression profile of CTS-N-induced papaya immunomodulatory response has not been reported. Here, the transcriptional map of papaya leaf genes were subjected to three treatments, viz., non-viral inoculation without CTS-N treatment (CK), virus inoculation without CTS-N treatment (CG), and virus inoculation of 1 g/L treatment (B). These were studied by pot culture experiment. Comparison of the B group with the CG group revealed 732 upregulated and 510 downregulated genes. Comparison of the CG group with the CK group revealed 909 upregulated and 1024 downregulated genes. To determine gene function, gene ontology (GO) analysis was performed, where 480 biological process genes, 256 molecular function genes, and 343 cell composition genes were differentially expressed. Kyoto Encyclopedia of Genes and Genomes (KEGG) results revealed that the top three pathways were phenylpropane biosynthesis, starch and sucrose metabolism, and plant hormone signal transduction. Real-time Quantitative PCR (qPCR) results were consistent with the transcriptome results, with a correlation coefficient of 0.87. The results of the transcriptional group showed that genes associated with plant resistance were induced by CTS-N-treatment in papaya. The chitinase gene was related to the plant disease process. Related genes in plant hormone signal transduction pathways are associated with plant resistance, and six differentially expressed genes were correlated with enhanced immune resistance in papaya.

MiR-301a promotes embryonic stem cell differentiation to cardiomyocytes.

BACKGROUND: Cardiovascular disease is the leading cause of death worldwide. Tissue repair after pathological injury in the heart remains a major challenge due to the limited regenerative ability of cardiomyocytes in adults. Stem cell-derived cardiomyocytes provide a promising source for the cell transplantation-based treatment of injured hearts. AIM: To explore the function and mechanisms of miR-301a in regulating cardiomyocyte differentiation of mouse embryonic stem (mES) cells, and provide experimental evidence for applying miR-301a to the cardiomyocyte differentiation induction from stem cells. METHODS: mES cells with or without overexpression of miR-301a were applied for all functional assays. The hanging drop technique was applied to form embryoid bodies from mES cells. Cardiac markers including GATA-4, TBX5, MEF2C, and α-actinin were used to determine cardiomyocyte differentiation from mES cells. RESULTS: High expression of miR-301a was detected in the heart from late embryonic to neonatal mice. Overexpression of miR-301a in mES cells significantly induced the expression of cardiac transcription factors, thereby promoting cardiomyocyte differentiation and beating cardiomyocyte clone formation. PTEN is a target gene of miR-301a in cardiomyocytes. PTEN-regulated PI3K-AKT-mTOR-Stat3 signaling showed involvement in regulating miR-301a-promoted cardiomyocyte differentiation from mES cells. CONCLUSION: MiR-301a is capable of promoting embryonic stem cell differentiation to cardiomyocytes.

Effective Neutralizing Antibody Produced in Mice Directly Immunized with Integrated Pichia pastoris Expressing HPV16L1 Protein.

The human papillomavirus (HPV) vaccine has not been widely used in developing countries because of its high cost and multiple subtype restrictions. The present study aimed to develop an economical, convenient, and effective vaccine to produce neutralizing antibodies. Using late protein 1 (L1) from the HPV16 subtype as the target antigen (HPV16L1) and Pichia pastoris as the antigen release system, integrated P. pastoris expressing HPV16L1 (named yeast-HPV16L1) was prepared and vaccinated directly into mice by subcutaneous multipoint injection. After immunization was performed thrice, high titers (greater than 1:40,960) of specific anti-HPV16L1 antibodies were obtained in immune serum and were observed to continuously rise over time. The indirect hemagglutination test and indirect hemagglutination inhibition test were used to detect neutralizing antibody activity in vitro, and the results demonstrated the hemagglutination ability of the immune serum and the reduction in or loss of the hemagglutination ability if preneutralized antigen was added to the immune serum. The protection conferred by immune serum to tumor-bearing mice at the early stages was confirmed, but the neutralizing activity disappeared when the tumor reached a size of 1 mm3. The neutralization activity of the immune serum was confirmed both in vitro and in vivo.

Study on Hesitant Fuzzy Information Measures and Their Clustering Application.

At present, research on hesitant fuzzy operations and measures is based on equal length processing, and an equal length processing method will inevitably destroy the original data structure and change the data information. This is an urgent problem to be solved in the development of hesitant fuzzy sets. Aiming at solving this problem, this paper firstly defines a hesitant fuzzy entropy function as the measure of the degree of uncertainty of hesitant fuzzy information and then proposes the concept of hesitant fuzzy information feature vector. The hesitant fuzzy distance measure and similarity measure are studied based on the information feature vector. Finally, the hesitant fuzzy network clustering method based on similarity measure is given, and the effectiveness of our algorithm through a numerical example is illustrated.

Expression analysis of four pseudo-response regulator (PRR) genes in Chrysanthemum morifolium under different photoperiods.

Genes encoding pseudo-response regulator (PRR) proteins play significant roles in plant circadian clocks. In this study, four genes related to flowering time were isolated from Chrysanthemum morifolium. Phylogenetic analysis showed that they are highly homologous to the counterparts of PRRs of Helianthus annuus and named as CmPRR2, CmPRR7, CmPRR37, and CmPRR73. Conserved motifs prediction indicated that most of the closely related members in the phylogenetic tree share common protein sequence motifs, suggesting functional similarities among the PRR proteins within the same subtree. In order to explore functions of the genes, we selected two Chrysanthemum varieties for comparison; that is, a short-day sensitive Zijiao and a short-day insensitive Aoyunbaixue. Compared to Aoyunbaixue, Zijiao needs 13 more days to complete the flower bud differentiation. Evidence from spatio-temporal gene expression patterns demonstrated that the CmPRRs are highly expressed in flower and stem tissues, with a growing trend across the Chrysanthemum developmental process. In addition, we also characterized the CmPRRs expression patterns and found that CmPRRs can maintain their circadian oscillation features to some extent under different photoperiod treatment conditions. These lines of evidence indicated that the four CmPRRs undergo circadian oscillation and possibly play roles in regulating the flowering time of C. morifolium.

Serum microRNA as potential biomarker to detect breast atypical hyperplasia and early-stage breast cancer.

AIM: The present study aimed to identify microRNA (miRNA) that can be used for not only detecting early-stage breast cancer (BC) but also diagnosing atypical hyperplasia (AH). MATERIALS & METHODS: RT-qPCR detected the expression levels of miRNAs and receiver operating characteristic curves were constructed to evaluate sensitivity and specificity of the assay. RESULTS: miR-24 and miR-103a were expressed in an upward trend in serum of benign proliferative tumor subjects, while they were downregulated significantly in serum of AH (p < 0.005) and early-stage BC subjects (p < 0.005) with high sensitivity and specificity as compared with controls. Bioinformatics analysis also revealed the potential molecular mechanism through which miR-24 and miR-103a regulate tumorigenesis in BC. CONCLUSION: miR-24 and miR-103a were valuable biomarkers for distinguishing AH and early-stage BC from healthy individuals/benign proliferative tumor patients.

Identification of a novel Hemoplasma species from pigs in Zhejiang province, China.

Hemoplasmas belong to Mycoplasmataceae (Mollicutes: Mycoplasmatales) and are able to infect a broad range of mammalian species. We investigated prevalence of hemotropic mycoplasma species in pig farms in the region of Zhejiang by a PCR scheme using universal primers targeting 16S rRNA and RNase P RNA gene (rnpB). Representative positive samples from different farms were selected for sequencing of 16S rRNA and the 219bp rnpB gene fragments for phylogenetic analysis. Sequencing analysis of PCR products from first samples identified a novel hemoplasma species present in several pig farms in the region with highest nucleotide identity of 92% to Candidatus Mycoplasma turicensis. A duplex PCR assay was then designed for differential detection of the novel hemoplasma from Mycoplasma parvum/M. suis in field samples. Of 324 blood samples from clinically healthy pigs, 26.5% was positive for this novel hemoplasma species and 50% positive for M. suis/M. parvum, indicating that the novel hemotropic mycoplasma species were of considerably high prevalence in Zhejiang province, China.

Major Immunodominant Region of Hepatitis B Virus Core Antigen as a Delivery Vector to Improve the Immunogenicity of the Fusion Antigen ROP2-SAG1 Multiepitope from Toxoplasma gondii in Mice.

To prepare the dominant multiepitope fusion antigen ROP2-SAG1 (RSmultiepitope) from Toxoplasma gondii in a prokaryotic system, the major immunodominant region (MIR) of the human hepatitis B virus core antigen (HBcAg(MIR)) was used as a delivery vector. The gene encoding the RSmultiepitope was inserted into HBcAg(MIR), and rHBcAg(MIR)-RSmultiepitope was prepared, purified, and administered to BALB/c mice through intradermal injection. An indirect enzyme-linked immunosorbent assay analysis based on a multiepitope peptide facilitated the specific differentiation of sera obtained from mice immunized with the rHBcAg(MIR)-RSmultiepitope protein, and high titers (greater than 1:6,400) of specific anti-RSmultiepitope antibodies were obtained. Immunized splenocytes demonstrated enhanced IFN-γ production. Based on these results, the HBcAg(MIR) vector is easily applied in vitro for targeting the RSmultiepitope and efficiently presents this target epitope for the induction of significant humoral and cellular immune responses. This study offers a novel strategy for the design of a target epitope delivery system for a toxoplasmosis vaccine.

An Empirical Assessment of Transgene Flow from a Bt Transgenic Poplar Plantation.

To assess the possible impact of transgenic poplar plantations on the ecosystem, we analyzed the frequency and distance of gene flow from a mature male transgenic Populus nigra plantation carrying the Bacillus thuringiensis toxin gene (Bt poplar) and the survival of Bt poplar seeds. The resultant Bt poplar seeds occurred at a frequency of ~0.15% at 0 m to ~0.02% at 500 m from the Bt poplar plantation. The germination of Bt poplar seeds diminished within three weeks in the field (germination rate from 68% to 0%) compared to 48% after three weeks of storage at 4°C. The survival rate of seedlings in the field was 0% without any treatment but increased to 1.7% under the addition of four treatments (cleaning and trimming, watering, weeding, and covering with plastic film to maintain moisture) after being seeded in the field for eight weeks. The results of this study indicate that gene flow originating from the Bt poplar plantation occurred at an extremely low level through pollen or seeds under natural conditions. This study provides first-hand field data on the extent of transgene flow in poplar plantations and offers guidance for the risk assessment of transgenic poplar plantations.