Defining ovine dermal papilla cell markers and identifying key signaling pathways regulating its intrinsic properties.

Dermal papilla cell (DPC), one of the key cell types during hair follicle development and regeneration, specifies hair size, shape and cycling. It is also an important in vitro screening model for hair growth. Although some characteristics of DPCs, such as agglutinative growth and marker genes, have been studied in mice and humans, the intrinsic properties of ovine DPCs and the regulatory mechanism of the intrinsic properties during continued culture in vitro remained unknown. In this study, based on our previous single-cell transcriptome sequencing on sheep lambskin, we verified SOX18 and PDGFRA as the novel marker genes of ovine DPCs through immunofluorescence staining on skin sections and cultured DPCs. Using continued cell culture and alkaline phosphatase staining, we found that different from mice and humans, ovine DPCs exhibit particularly robust and stable aggregation with unbated alkaline phosphatase activity till 30 passages during continued culture in vitro. Also, we found that the expression of some marker genes and the activity of Wnt/ß-catenin signaling differ between early passaged DPCs and multiple passaged DPCs. Further, using Wnt/ß-catenin agonist and antagonist, we demonstrated that Wnt/ß-catenin signaling could regulate cell aggregation and alkaline phosphatase activity of ovine DPCs through regulating FGF and IGF signaling. This study provides the basis for isolating ovine DPCs and defines their intrinsic properties, which contribute to improving wool performance and medicine of hair regeneration.

Preliminary study of melatonin in the proliferation and apoptosis of Hu sheep dermal papilla cells in vitro.

Previous studies have shown that melatonin has a certain regulatory effect on the growth of sheep wool. However, the mechanism of melatonin action remains unknown. In the present study, we aimed to understand the role of exogenous melatonin in the dermal papilla cells of Hu sheep. To confirm the optimal melatonin treatment regimen for Hu sheep dermal papilla cells, we detected the cell viability by exposing them to different concentrations of melatonin and different treatment times. The results showed that cell viability was best when dermal papilla cells were treated with 1000 pg/ml of melatonin for 48 h. According to the results of qPCR, CCK-8, EDU, Western blot, and Flow cytometry analysis, we found that 1000 pg/ml melatonin promoted the proliferation and inhibited the apoptosis of dermal papilla cells compared with the exogenous melatonin blank group (control group). Furthermore, we also found that 1000 pg/ml of melatonin promoted the cell cycle progress of dermal papilla cells according to the results of qPCR and Flow cytometry analysis. Overall, our findings showed that melatonin plays an important role in the dermal papilla cells of Hu sheep.

Insights Into Long Non-Coding RNA and mRNA Expression in the Jejunum of Lambs Challenged With Escherichia coli F17.

It has long been recognized that enterotoxigenic Escherichia coli (ETEC) is the major pathogen responsible for vomiting and diarrhea. E. coli F17, a main subtype of ETEC, is characterized by high morbidity and mortality in young livestock. However, the transcriptomic basis underlying E. coli F17 infection has not been fully understood. In the present study, RNA sequencing was conducted to explore the expression profiles of mRNAs and long non-coding RNAs (lncRNAs) in the jejunum of lambs who were identified as resistant or sensitive to E. coli F17 that was obtained in a challenge experiment. A total of 772 differentially expressed (DE) mRNAs and 190 DE lncRNAs were detected between the E. coli F17-resistance and E. coli F17-sensitive lambs (i.e., TFF2, LOC105606142, OLFM4, LYPD8, REG4, APOA4, TCONS_00223467, and TCONS_00241897). Then, a two-step machine learning approach (RX) combination Random Forest and Extreme Gradient Boosting were performed, which identified 16 mRNAs and 17 lncRNAs as potential biomarkers, within which PPP2R3A and TCONS_00182693 were prioritized as key biomarkers involved in E. coli F17 infection. Furthermore, functional enrichment analysis showed that peroxisome proliferator-activated receptor (PPAR) pathway was significantly enriched in response to E. coli F17 infection. Our finding will help to improve the knowledge of the mechanisms underlying E. coli F17 infection and may provide novel targets for future treatment of E. coli F17 infection.

The effect of quercetin on diabetic nephropathy (DN): a systematic review and meta-analysis of animal studies.

Quercetin, a flavonoid possessing numerous biological activities, is reported to improve renal injury in diabetic animals. Here, the aim of this systematic review and meta-analysis is to assess the effect of quercetin on diabetic nephropathy and summarize its possible mechanisms. We searched in four databases PubMed, Web of Sciences (WOS), Cochrane and Embase from inception to May 2021 and ultimately included 20 animal studies in this review. A total of 12 outcome measurements including renal function indexes, oxidative stress biomarkers and inflammatory cytokines were extracted for meta-analysis using RevMan 5.4 software. Apart from creatinine clearance and uric acid with no significant difference, quercetin significantly decreased the levels of renal index, serum/plasma creatinine (SCr), blood urea nitrogen (BUN), urine protein, urine albumin, malondialdehyde (MDA), tumor necrosis factor (TNF)-α and interleukin (IL)-1ß, and increased superoxide dismutase (SOD) and catalase (CAT) activity. In short, quercetin improves renal function and attenuates the renal oxidative stress level and inflammatory response in DN animal models. Its possible action mechanisms include anti-oxidation, anti-inflammation, anti-fibrosis, and regulation of renal lipid accumulation.

Dietary phenolic-type Nrf2-activators: implications in the control of toxin-induced hepatic disorders.

Numerous studies have exemplified the importance of nuclear factor erythroid 2-related factor 2 (Nrf2) activation in the alleviation of toxin-induced hepatic disorders primarily through eliminating oxidative stress. Whereafter, increasingly more efforts have been contributed to finding Nrf2-activators, especially from dietary polyphenols. The present review summarized the phenolic-type Nrf2-activators published in the past few decades, analyzed their effectiveness based on their structural characteristics and outlined their related mechanisms. It turns out that flavonoids are the largest group of phenolic-type Nrf2-activators, followed by nonflavonoids and phenolic acids. When counting on subgroups, the top three types are flavonols, flavones, and hydroxycinnamic acids, with curcuminoids having the highest effective doses. Moreover, most polyphenols work through the phosphorylation of Nrf2. Besides, mitogen-activated protein kinases (MAPKs) and protein kinase B (Akt) are the frequent targets of these Nrf2-activators, which indirectly mediate the behavior of Nrf2. However, current data are not sufficient to conclude any structure-activity relationship.

Non-Coding Transcriptome Provides Novel Insights into the Escherichia coli F17 Susceptibility of Sheep Lamb.

It has long been recognized that enterotoxigenic Escherichia coli (ETEC) is the major pathogen responsible for vomiting and diarrhea. E. coli F17, a main subtype of ETEC, is characterized by high morbidity and mortality in young livestock. However, the transcriptomic basis underlying E. coli F17 infection has not been fully understood. In this study, RNA sequencing was performed to explore the expression profiles of circRNAs and miRNAs in the jejunum of E. coli F17-antagonism (AN) and -sensitive (SE) lambs. A total of 16,534 circRNAs and 271 miRNAs (125 novel miRNAs and 146 annotated miRNAs) were screened, and 214 differentially expressed (DE) circRNAs and 53 DE miRNAs were detected between the AN and SE lambs (i.e., novel_circ_0025840, novel_circ_0022779, novel_miR_107, miR-10b). Functional enrichment analyses showed that source genes of DE circRNAs were mainly involved in metabolic-related pathways, while target genes of DE miRNAs were mainly enriched in the immune response pathways. Then, a two-step machine learning approach combining Random Forest (RF) and XGBoost (candidates were first selected by RF and further assessed by XGBoost) was performed, which identified 44 circRNAs and 39 miRNAs as potential biomarkers (i.e., novel_circ_0000180, novel_circ_0000365, novel_miR_192, oar-miR-496-3p) for E. coli infection. Furthermore, circRNA-related and lncRNA-related ceRNA networks were constructed, containing 46 circRNA-miRNA-mRNA competing triplets and 630 lncRNA-miRNA-mRNA competing triplets, respectively. By conducting a serious of bioinformatic analyses, our results revealed important circRNAs and miRNAs that could be potentially developed as candidate biomarkers for intestinal inflammatory response against E. coli F17 infection; our study can provide novel insights into the underlying mechanisms of intestinal immunity.

Nitric Oxide and Immune Responses in Cancer: Searching for New Therapeutic Strategies.

In recent years, there has been an increasing interest in understanding the mysterious functions of nitric oxide (NO) and how this pleiotropic signaling molecule contributes to tumorigenesis. This review attempts to expose and discuss the information available on the immunomodulatory role of NO in cancer and recent approaches to the role of NO donors in the area of immunotherapy. To address the goal, the following databases were searched to identify relevant literature concerning empirical evidence: The Cochrane Library, Pubmed, Medline, and EMBASE from 1980 through March 2020. Valuable attempts have been made to develop distinctive NO-based cancer therapy. Although the data do not allow generalization, the evidence seems to indicate that low/moderate levels may favor tumorigenesis, while higher levels would exert antitumor effects. In this sense, the use of NO donors could have an important therapeutic potential within immunotherapy, although there are still no clinical trials. The emerging understanding of NO-regulated immune responses in cancer may help unravel the recent features of this "doubleedged sword" in cancer physiological and pathologic processes and its potential use as a therapeutic agent for cancer treatment. In short, in this review, we discuss the complex cellular mechanism in which NO, as a pleiotropic signaling molecule, participates in cancer pathophysiology. We also debate the dual role of NO in cancer and tumor progression and clinical approaches for inducible nitric oxide synthase (iNOS) based therapy against cancer.

Effect of Sox18 on the Induction Ability of Dermal Papilla Cells in Hu Sheep.

Sox18 is a developmental gene that encodes transcription factors. It has been indicated as be a key gene affecting the growth and development of hair follicles, in which dermal papilla cells (DPCs) have been demonstrated to play an important role through their ability to induce the formation of hair follicles. Pre-laboratory studies have found that Sox18 is differentially expressed in the dermal papilla cells of different pattern types of Hu sheep. We speculated that Sox18 plays an important role in the dermal papilla cells of Hu sheep. In our study, we analyzed the effect of Sox18 on the induction ability of DPCs in order to elucidate the function and molecular mechanism of Sox18 in the DPCs of Hu sheep. We first identified the expression of Sox18 in the DPCs of Hu sheep by immunofluorescence staining. We then used alkaline phosphatase staining, cell morphology observations and RT-PCR to detect the effect of Sox18 on the induction of DPCs after overexpression of or interference with Sox18. We also used RT-PCR, WB and immunofluorescence staining to detect the effect of Sox18 on the Wnt/ß-catenin signal pathway in DPCs. We found that Sox18 was specifically expressed in the DPCs of Hu sheep, and that Sox18 could enhance the alkaline phosphatase activity in the DPCs of Hu sheep and accelerate cell agglutination. The results of RT-PCR revealed that Sox18 promoted the mRNA expression of Versican, HHIP and FGFRI, and inhibited the mRNA expression of BMP4 and WIF1. Further studies showed that Sox18 promoted the expression of ß-catenin and activated the Wnt/ß-catenin signal pathway in DPCs. When the Wnt/ß-catenin signal pathway of DPCs was activated, the induction ability of DPCs was enhanced. Overall, we believe that Sox18 could enhance the induction ability of DPCs in Hu sheep and regulate the induction ability of DPCs through the Wnt/ß-catenin signal pathway.

miR-143 Targeting CUX1 to Regulate Proliferation of Dermal Papilla Cells in Hu Sheep.

Wool curvature is the determining factor for lambskin quality of Hu lambs. However, the molecular mechanism of wool curvature formation is not yet known. miRNA has been proved to play an important role in hair follicle development, and we have discovered a differentially expressed miRNA, miR-143, in hair follicles of different curl levels. In this study, we first examined the effects of miR-143 on the proliferation and cell cycle of dermal papilla cells using CCK8, EdU and flow cytometry and showed that miR-143 inhibited the proliferation of dermal papilla cells and slowed down the cell cycle. Bioinformatics analysis was performed to predict the target genes KRT71 and CUX1 of miR-143, and both two genes were expressed at significantly higher levels in small waves than in straight lambskin wool (p < 0.05) as detected by qPCR and Western blot (WB). Then, the target relationships between miR-143 and KRT71 and CUX1 were verified through the dual-luciferase assay in 293T cells. Finally, after overexpression and suppression of miR-143 in dermal papilla cells, the expression trend of CUX1 was contrary to that of miR-143. Meanwhile, KRT71 was not detected because KRT71 was not expressed in dermal papilla cells. Therefore, we speculated that miR-143 can target CUX1 to inhibit the proliferation of dermal papilla cells, while miR-143 can target KRT71 to regulate the growth and development of hair follicles, so as to affect the development of hair follicles and ultimately affect the formation of wool curvature.

EZH2 Exacerbates Breast Cancer by Methylating and Activating STAT3 Directly.

Breast cancer is one of the most common causes of female death globally. Numerous clinical drugs for breast cancer have been developed, but the unsatisfactory, inevitable side effects and drug resistance are the emerging threatens. Therefore, it is necessary to investigate the comprehensive mechanism of breast cancer. Enhancer of zeste homolog 2 (EZH2) is a candidate oncogenic driver in diverse cancers, such as breast cancer. The canonical role of EZH2 has been vastly investigated, but the non-canonical function of EZH2 in breast cancer remains unclear. Here, we demonstrated that EZH2 exacerbated breast cancer in non-canonical manner by methylating STAT3. EZH2 over-expressed in breast cancer patients and regulated STAT3 post-transcriptionally according to TCGA datasets. Chemical and genetic inhibition of EZH2 impeded proliferation and migration of breast cancer cells, which may be partially rescued by STAT3 over-expression. EZH2 physically interacted with STAT3 and methylated STAT3 directly, resulting in increased nuclear localization and chromatin of STAT3. Furthermore, the mutation of STAT3 methylation site, targeted by EZH2, impeded the transcriptional activity of STAT3. Eventually, disturbed STAT3 methylation by EZH2 in animal model showed decreased breast cancer growth. These data confirm that EZH2 exacerbates breast cancer by methylating STAT3 directly, and thus providing a promising therapeutic target for breast cancer.

Flaxseed extract induces apoptosis in human breast cancer MCF-7 cells.

Significant evidence indicated that flaxseed (Linum usitatissimum) possesses various positive health aspects such as reducing the risk of cancer and cardiovascular diseases. The fatty acids are considered to be responsible for these benefits of flaxseed. Herein, the in vitro effects of flaxseed extract on the growth and apoptosis of human breast cancer MCF-7 cells were investigated. The MCF-7 cells treated with flaxseed extract showed a dose-dependent decrease in cell viability. The flaxseed extract induced reactive oxygen species and the flow cytometric analysis demonstrated that flaxseed fatty acids triggered apoptosis of MCF-7 cells, which was also shown by the loss of mitochondrial membrane potential and caspase cascade reaction. Thus, the flaxseed extract regulated the growth of MCF-7 cells and induced apoptosis. Eventually, the flaxseed could be used as a dietary supplement to prevent breast cancer.