Fluorescence detection of apolipoprotein E gene polymorphisms based on oligonucleotide ligation and magnetic separation.

Alzheimer's disease is a progressive neurodegenerative condition that causes brain cell death and is the leading cause of dementia. Most patients with Alzheimer's disease are diagnosed with late-onset Alzheimer's disease (LOAD), with apolipoprotein E (APOE) genotypes being highly associated with the frequency of LOAD risk. A fluorescence detection system coupled with oligonucleotide ligation and magnetic separation was developed to identify two single-nucleotide polymorphisms (SNPs) for the APOE gene and recognize APOE alleles for LOAD. The system utilized a fluorescence probe with one base-discriminating nucleoside for SNP (F probe) and a perfectly complementary biotin-modified sequence against the target DNA (P probe). When the F and P probes matched the target DNA sequences, DNA ligation occurred, and ligation products were produced. Streptavidin magnetic beads were subsequently employed to remove the ligation products, and a decrease in fluorescence intensity was observed in the supernatant compared to when there was no target DNA. This system detected two SNPs of APOE alleles, namely rs429358 and rs7412. The results indicated that the R-values ((F0 - F1)/F0) for rs429358 were 0.92 ± 0.002 for the T/T target, 0.47 ± 0.004 for the T/C target and 0.11 ± 0.004 for the C/C target, respectively. The R-values for rs7412 were 0.73 ± 0.009 for the C/C target, 0.42 ± 0.001 for the C/T target and 0.16 ± 0.007 for the T/T target, respectively. F0 and F1 represent the fluorescence intensity of the F probe without and with target DNA, respectively. Based on fluorescence intensity, the fluorescence detection system was able to identify the genotypes of the APOE gene accurately to evaluate the risk of Alzheimer's disease.

Lipid Accumulation in Blastocystis Increases Cell Damage in Co-Cultured Cells.

Blastocystis hominis is an intestinal protozoan that is often neglected, despite causing abdominal pain and diarrhea. Previous research has demonstrated that lipids can be synthesized by B. hominis or can accumulate in growth medium, but their function and mechanisms in the pathogenesis of Blastocystis remain unclear. Our study found that lipid-rich Blastocystis ST7-B can increase inflammation and disrupt Caco-2 cells more than the same parasite without the lipovenoes supplement. Additionally, the cysteine protease of Blastocystis, a virulence factor, is upregulated and has higher activity in lipid-rich Blastocystis. In order to better understand the effects of lipids on Blastocystis pathogenesis, we treated lipid-lowering pravastatin during Blastocystis ST7-B culturing with a lipovenoes supplement, which decreased the lipid levels of the Blastocystis and reduced the Blastocystis-induced inflammation and cell disruption of Caco-2 cells. We also analyzed the fatty acid composition and possible synthesis pathway in Blastocystis ST7-B, finding significantly higher ratios of arachidonic acid, oleic acid, and palmitic acid than in the other lipid components in lipid-rich Blastocystis ST7-B. These results suggest that lipids play a significant role in the pathogenesis of Blastocystis and provide important information on the molecular mechanisms of and potential treatments for Blastocystis infection.

The Role of Autophagy in Anti-Cancer and Health Promoting Effects of Cordycepin.

Cordycepin is an adenosine derivative isolated from Cordyceps sinensis, which has been used as an herbal complementary and alternative medicine with various biological activities. The general anti-cancer mechanisms of cordycepin are regulated by the adenosine A3 receptor, epidermal growth factor receptor (EGFR), mitogen-activated protein kinases (MAPKs), and glycogen synthase kinase (GSK)-3ß, leading to cell cycle arrest or apoptosis. Notably, cordycepin also induces autophagy to trigger cell death, inhibits tumor metastasis, and modulates the immune system. Since the dysregulation of autophagy is associated with cancers and neuron, immune, and kidney diseases, cordycepin is considered an alternative treatment because of the involvement of cordycepin in autophagic signaling. However, the profound mechanism of autophagy induction by cordycepin has never been reviewed in detail. Therefore, in this article, we reviewed the anti-cancer and health-promoting effects of cordycepin in the neurons, kidneys, and the immune system through diverse mechanisms, including autophagy induction. We also suggest that formulation changes for cordycepin could enhance its bioactivity and bioavailability and lower its toxicity for future applications. A comprehensive understanding of the autophagy mechanism would provide novel mechanistic insight into the anti-cancer and health-promoting effects of cordycepin.

Magnoliae Flos Essential Oil as an Immunosuppressant in Dendritic Cell Activation and Contact Hypersensitivity Responses.

Magnoliae Flos is a commonly used traditional medicinal material in Asia. It is used to treat sinusitis, nasal congestion, and hypersensitive skin. Because Magonlia Flos was described as an aromatic material in ancient Chinese texts, we hypothesized that its essential oil may be used to treat immune disorders. Dendritic cells (DCs), regarded as a major target of immunomodulators to control immune responses, play a critical role in the adaptive immune response. In this study, Magnoliae Flos essential oil (MFEO) decreased the production of the cytokines TNF-α, IL-6, and IL-12p70 in lipopolysaccharide (LPS)-stimulated DCs. It also suppressed the surface markers MHC II, CD80, and CD86 in LPS-stimulated DCs. Animal models demonstrated that the 2,4-Dinitro-1-fluorobenzene (DNFB) inducing a contact hypersensitivity response was inhibited following treatment with MFEO. In addition, MFEO inhibited the infiltration of T cells in the ears of DNFB-induced mice. To explore its bioactive compounds, the components of MFEO were analyzed using gas chromatography (GC) and GC-mass spectrometry. The results revealed that the major compounds in MFEO are camphor and 1,8-cineole. Additional DC bioassays confirmed that these compounds substantially suppressed cytokine production in LPS-induced DCs. Therefore, we demonstrated that MFEO exhibits an immunosuppressive effect both in vivo and in vitro, and camphor and 1,8-cineole may be the major components responsible for its immunosuppressive ability. The findings indicate that MFEO has the potential to be developed as a new immunosuppressant for excessive diseases.

Fire-heat and Qi deficiency syndromes as predictors of short-term prognosis of acute ischemic stroke.

OBJECTIVES: To explore the relationships between traditional Chinese medicine (TCM) syndromes and disease severity and prognoses after ischemic stroke, such as neurologic deficits and decline in activities of daily living (ADLs). METHODS: The study included 211 patients who met the inclusion criteria of acute ischemic stroke based on clinical manifestations, computed tomography or magnetic resonance imaging findings, and onset of ischemic stroke within 72 hours with clear consciousness. To assess neurologic function and ADLs in patients with different TCM syndromes, the TCM Syndrome Differentiation Diagnostic Criteria for Apoplexy scale (containing assessments of wind, phlegm, blood stasis, fire-heat, qi deficiency, and yin deficiency with yang hyperactivity syndromes) was used within 72 hours of stroke onset, and Western medicine-based National Institutes of Health Stroke Scale (NIHSS) and Barthel Index (BI) assessments were performed at both admission and discharge. RESULTS: The most frequent TCM syndromes associated with acute ischemic stroke were wind syndrome, phlegm syndrome, and blood stasis syndrome. Improvement according to the BI at discharge and days of admission were significantly different between patients with and those without fire-heat syndrome. Patients with qi deficiency syndrome had longer hospital stays and worse NIHSS and BI assessments at discharge than patients without qi deficiency syndrome. All the reported differences reached statistical significance. CONCLUSIONS: These results provide evidence that fire-heat syndrome and qi deficiency syndrome are essential elements that can predict short-term prognosis of acute ischemic stroke.

Areca (betel) nut extract activates mitogen-activated protein kinases and NF-kappaB in oral keratinocytes.

Areca (betel) was recently proved a carcinogenic substance by the International Agency for Research on Cancer. However, the signaling impact of areca in oral keratinocyte is still obscure. Mitogen-activated protein kinase superfamilies, including extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinases (JNK) and p38, together with transcription factor NF-kappaB, are important signaling elements. We examined the activation of these signaling pathways in OECM-1 and SAS oral keratinocytes, treated with ripe areca nut extract (ANE). In both cells, a rapid increase in JNK1 activity at 0.5 hr was noted following treatment of ANE. ERK was profoundly activated during 0.5-2 hr in OECM-1 cells. Contrasting p38 activity was noted in these 2 cells. In both cells, ANE also activated NF-kappaB pathway in a biphasic manner, particularly for SAS cells. NF-kappaB was activated by approximately 2- to 4-fold at 0.5-1 hr and a plateau or slight decrease of activity existed between 1 and 6 hr. Later, another higher episode of NF-kappaB activity was raised. This was accompanied with the rapid degradation in cytosolic IkappaBalpha as well as an increase of nuclear NF-kappaB in both cells. ANE treatment did not activate epidermal growth factor receptor signaling system, but blockage of NF-kappaB activation rendered the suppression of ANE-modulated COX-2 upregulation in OECM-1. This study identified that ANE affected interactive signaling systems in oral keratonocytes that could be the pathogenetic basis for areca.