Experimental Biology and Medicine

Intervertebral disc degeneration is a complex disease involving genetic and environmental factors and multiple cellular processes. The role and expression of the lncRNA NEAT1 were assessed in intervertebral disc degeneration. NEAT1 expression was assessed in degenerative and control nucleus pulposus using RT-PCR. Western blotting and RT-PCR were also used to investigate p53 and p21 levels in nucleus pulposus tissues. NEAT1 function in degenerative nucleus pulposus cells was assessed with gain- and loss-of-function experiments. ERK/MAPK signaling was also examined. NEAT1, p53, and p21 were dramatically upregulated in intervertebral disc degeneration. Furthermore, catabolic MMP13 and ADAMTS5 were dysregulated and collagen II and aggrecan were downregulated after NEAT1 overexpression. This effect was reversed by transfection with si-NEAT1 in degenerative nucleus pulposus cells. In addition, NEAT1 was found to affect the activation of the ERK/MAPK pathway. The NEAT1-induced ECM degradation may involve ERK1/2/MAPK signaling. LncRNA NEAT1 may represent a novel molecular target for intervertebral disc degeneration treatment by preventing nucleus pulposus ECM degradation.

Cardiac fibroblasts and their activated derivatives, myofibroblasts, play a critical role in wound healing after myocardial injury and often contribute to long-term pathological outcomes, such as excessive fibrosis. Thus, defining the microenvironmental factors that regulate the phenotype of cardiac fibroblasts and myofibroblasts could lead to new therapeutic strategies. Both chemical and biomechanical cues have previously been shown to induce myofibroblast differentiation in many organs and species. For example, transforming growth factor beta 1, a cytokine secreted by neutrophils, and rigid extracellular matrix environments have both been shown to promote differentiation. However, the relative contributions of transforming growth factor beta 1 and extracellular matrix rigidity, two hallmark cues in many pathological myocardial microenvironments, to the phenotype of human cardiac fibroblasts are unclear. We hypothesized that transforming growth factor beta 1 and rigid extracellular matrix environments would potentially have a synergistic effect on the differentiation of human cardiac fibroblasts to myofibroblasts. To test this, we seeded primary human adult cardiac fibroblasts onto coverslips coated with polydimethylsiloxane of various elastic moduli, introduced transforming growth factor beta 1, and longitudinally quantified cell phenotype by measuring expression of α-smooth muscle actin, the most robust indicator of myofibroblasts. Our data indicate that, although extracellular matrix rigidity influenced differentiation after one day of transforming growth factor beta 1 treatment, ultimately transforming growth factor beta 1 superseded extracellular matrix rigidity as the primary regulator of myofibroblast differentiation. We also measured expression of POSTN, FAP, and FSP1, proposed secondary indicators of fibroblast/myofibroblast phenotypes. Although these genes partially trended with α-smooth muscle actin expression, they were relatively inconsistent. Finally, we demonstrated that activated myofibroblasts incompletely revert to a fibroblast phenotype after they are re-plated onto new surfaces without transforming growth factor beta 1, suggesting differentiation is partially reversible. Our results provide new insights into how microenvironmental cues affect human cardiac fibroblast differentiation in the context of myocardial pathology, which is important for identifying effective therapeutic targets and dictating supporting cell phenotypes for engineered human cardiac disease models.

Dysbiosis, a broad spectrum of imbalance of the gut microbiota, may progress to microbiota dysfunction. Dysbiosis is linked to some human diseases, such as inflammation-related disorders and metabolic syndromes. However, the underlying mechanisms of the pathogenesis of dysbiosis remain elusive. Recent findings suggest that the microbiome and gut immune responses, like immunoglobulin A production, play critical roles in the gut homeostasis and function, and the progression of dysbiosis. In the past two decades, much progress has been made in better understanding of production of immunoglobulin A and its association with commensal microbiota. The present minireview summarizes the recent findings in the gut microbiota dysbiosis and dysfunction of immunoglobulin A induced by the imbalance of pathogenic bacteria and commensal microbiota. We also propose the potentials of dietary carotenoids, such as β-carotene and astaxanthin, in the improvement of the gut immune system maturation and immunoglobulin A production, and the consequent promotion of the gut health.

A major characteristic of immunodeficiency associated with life-threatening intracellular infection in adults is the presence of anti-interferon-γ antibodies. Although little is known about the mechanism underlying this syndrome, it is believed that the antibodies inhibit the activity of downstream signaling pathway of interferon-γ. In this study, the characteristics of these antibodies in patients who presented, or have a history of, intracellular infection and were positive to anti-interferon-γ antibodies were investigated. The antibodies exhibited mainly the IgG1 and the IgG4 subtypes and recognized the C-terminal of the interferon-γ linear epitope containing the KRKR motif, which is required for the biological activity of interferon-γ. The antibodies bound to recombinant interferon-γ with significantly lower avidity than antibodies to a recall antigen, tetanus toxoid, suggesting that the antibodies might have not undergone affinity maturation. The data from this study may provide fundamental information to better understand the properties of anti-interferon-γ antibodies, which can be useful for future studies.

A defining feature of neurodegenerative diseases is the abnormal and excessive loss of neurons. One molecule that is particularly important in promoting neuronal death in a variety of cell culture and in vivo models of neurodegeneration is histone deacetylase-3 (HDAC3), a member of the histone deacetylase family of proteins. As a step towards understanding how HDAC3 promotes neuronal death, we conducted a proteomic screen aimed at identifying proteins that were regulated by HDAC3. HDAC3 was overexpressed in cultured rat cerebellar granule neurons (CGNs) and protein lysates were analyzed by mass spectrometry. Of over 3000 proteins identified in the screen, only 21 proteins displayed a significant alteration in expression. Of these, 12 proteins were downregulated whereas 9 proteins were upregulated. The altered expression of five of these proteins, TEX10, NPTX1, TFG, TSC1, and NFL, along with another protein that was downregulated in the proteomic screen, HIP1R, was confirmed using Western blots and commercially available antibodies. Because antibodies were not available for some of the proteins and since HDAC3 is a transcriptional regulator of gene expression, we conducted RT-PCR analysis to confirm expression changes. In separate analyses, we also included other proteins that are known to regulate neurodegeneration, including HDAC9, HSF1, huntingtin, GAPDH, FUS, and p65/RELA. Based on our proteomic screen and candidate protein approach, we identify three genes, Nptx1, Hip1r, and Hdac9, all known to regulate neurodegeneration that are robustly regulated by HDAC3. Given their suggested roles in regulating neuronal death, these genes are likely to be involved in regulating HDAC3-mediated neurotoxicity.

Rhabdomyolysis (destruction of striated muscle) is a novel form of mushroom poisoning in Europe and Asia indicated by increased circulating creatine kinase levels. Particular wild fungi have also been reported to induce elevated creatine kinase activities in mice. Flammulina velutipes (enokitake or winter mushroom) is one of the most actively cultivated mushroom species globally. As it is marketed as a medicinal mushroom and functional food, it is important to examine whether it could induce potentially harmful health effects similar to some previously studied edible fungi. The present study examined the effects of F. velutipes consumption on the plasma clinical chemistry, hematology, and organ histology of laboratory mice. Wild F. velutipes were dried, pulverized, mixed with a regular laboratory rodent diet, and fed to the animals at 0, 3, 6, or 9 g/kg body mass/day for five days (n = 6/group). F. velutipes consumption caused increased activities of plasma creatine kinase and the MB-fraction of creatine kinase at 6–9 g/kg/d, indicating potentially deleterious effects on both skeletal and cardiac muscle. The plasma total and high-density lipoprotein cholesterol concentrations (at 9 g/kg/d) and white blood cell and lymphocyte counts (at 6–9 g/kg/d) decreased. Although the cholesterol-lowering properties of F. velutipes can be beneficial, the previously unexamined, potentially hazardous side effects of mushroom consumption (myo- and cardiotoxicity) should be thoroughly investigated before recommending this mushroom species as a health-promoting food item.

Hepatocellular carcinoma (HCC) is one of the most malignant diseases worldwide. The unfavorable clinical outcome and poor prognosis are due to high rates of recurrence and metastasis after treatments. Some scholars of traditional Chinese medicine suggested that endogenous wind-evil had played an important role in metastasis of malignant tumor. Therefore, the drug of dispelling wind-evil could be used to prevent cancer metastasis and improve the poor prognosis. So we wondered whether Scorpion, one of the most important wind calming drugs, has antitumor effect especially in epithelial–mesenchymal transition (EMT) and metastasis of HCC in this research. We found that Scorpion-medicated serum could inhibit proliferation, induce apoptosis, and decrease migration and invasion capacity of Hepa1-6 cells in vitro. Meanwhile, we observed that water decoction of Scorpion restrained tumor growth and metastasis in nude mouse of HCC metastasis models. Further experiments showed that Scorpion could suppress EMT, which is characterized by increased epithelial marker E-cadherin expression and decreased mesenchymal markers N-cadherin and Snail expression following Scorpion treatment both in vitro and in vivo. These results suggested that the Scorpion could inhibit Hepa1-6 cells’ invasion and metastasis in part by reversing EMT and providing a possible potential approach for preventing HCC metastasis.

This study is to investigate transcription factors involved in cisplatin resistance in ovarian cancer cells. The transcriptome of cisplatin resistant and sensitive A2780 epithelial ovarian cancer cells was obtained from GSE15372. Ovarian transcriptome data GSE62944 was downloaded from TCGA and applied for transcription regulatory network analysis. The analysis results were confirmed using quantitative polymerase chain reaction. The roles of SREBP2 in cisplatin-resistant cells were investigated by RNA inference and cell viability analysis. Transcription regulatory network analysis found that 12 transcription factors and their targets were involved in cisplatin resistant in A2780 cells. Among these factors, the targets of EZH2 and SREBP2 revealed by Transcriptional Regulatory Relationships Unraveled by Sentence-based Text mining were also enriched in differentially expressed genes between cisplatin resistant and cisplatin sensitive cells. Their targets were enriched mainly in cell cycle and cholesterol metabolic process, respectively. Bioinformatic analysis illustrated three known targets of SREBP2, namely LDLR, FDFT1, and HMGCR were increased in A2780-resistant cell lines. Additionally, the three genes and SREBP2 were also elevated in live cells after cisplatin treatment via quantitative polymerase chain reaction. Importantly, RNA inference of SREBP2 in A2780 cell line resulted in a decrease of cell viability after cisplatin treatment. SREBP2 played important roles in cisplatin resistance and cholesterol metabolic process might be a novel target for cancer therapy.