Age-dependent effects of acute stress on the behavior, blood parameters, immunity, and enteric nerves of mice.

The impact of stress on mental and digestive health has been extensively studied, with chronic stress being associated with various disorders. However, age-related differences in the response to acute stress, both behaviorally and physiologically, remain poorly understood. Therefore, this study aimed to develop a model to detect transient stress in mice of different ages. The stressor employed in our experiments was a restraint stress procedure, where mice were subjected to brief periods of immobilization to induce an acute stress response. Male C3H/HeN mice aged 3, 6, 12, and 30 weeks were subjected to acute restrain stress (ARS) by being placed in a 50 ml conical centrifuge tube for 15 min. Subsequently, their behavior, organ tissues, hematological parameters, cortisol concentration, and immune responses were assessed. Following ARS, the increased in time and entries into the center by the 12-week-old mice following stress. In comparison to mice of other ages, those aged 6 weeks demonstrated notable elevations in erythrocytes, platelets, hemoglobin, and hematocrit, all of which were influenced by the time-dependent changes and the recovery process of ARS. Blood corticosterone levels were substantially elevated in all age groups after ARS. Furthermore, ARS induced a notable increase in leukocytes, basophils, residential macrophages, and CD4+ T cells in all age groups except for 3-week-old mice. However, the number of monocyte-derived macrophages and CD8+ T cells did not change significantly. Additionally, mice aged 3 and 6 weeks demonstrated an increase in GFAP+ cells following ARS, whereas NeuN+ cells decreased across all ages. These results suggest that ARS has varying effects on the behavior, cortisol concentration, and quantity of blood cells as well as hepatic immune cells in mice of different ages. These age-dependent responses shed light on the complex interplay between stress and physiological systems and contribute to the broader understanding of stress-related diseases.

Extracellular domain of semaphorin 5A serves a tumor­suppressing role by activating interferon signaling pathways in lung adenocarcinoma cells.

Semaphorin 5A (SEMA5A), which was originally identified as an axon guidance molecule in the nervous system, has been subsequently identified as a prognostic biomarker for lung cancer in nonsmoking women. SEMA5A acts as a tumor suppressor by inhibiting the proliferation and migration of lung cancer cells. However, the regulatory mechanism of SEMA5A is not clear. Therefore, the purpose of the present study was to explore the roles of different domains of SEMA5A in its tumor­suppressive effects in lung adenocarcinoma cell lines. First, it was revealed that overexpression of full length SEMA5A or its extracellular domain significantly inhibited the proliferation and migration of both A549 and H1299 cells using MTT, colony formation and gap closure assays. Next, microarray analyses were performed to identify genes regulated by different domains of SEMA5A. Among the differentially expressed genes, the most significant function of these genes that were enriched was the 'Interferon Signaling' pathway according to Ingenuity Pathway Analysis. The activation of the 'Interferon Signaling' pathway was validated by reverse transcription­quantitative PCR and western blotting. In summary, the present study demonstrated that the extracellular domain of SEMA5A could upregulate genes in interferon signaling pathways, resulting in suppressive effects in lung adenocarcinoma cells.

Indoxyl Sulfate Elevated Lnc-SLC15A1-1 Upregulating CXCL10/CXCL8 Expression in High-Glucose Endothelial Cells by Sponging MicroRNAs.

Cardiovascular disease (CVD) is the leading cause of mortality in diabetes mellitus (DM). Immunomodulatory dysfunction is a primary feature of DM, and the emergence of chronic kidney disease (CKD) in DM abruptly increases CVD mortality compared with DM alone. Endothelial injury and the accumulation of uremic toxins in the blood of DM/CKD patients are known mechanisms for the pathogenesis of CVD. However, the molecular factors that cause this disproportional increase in CVD in the DM/CKD population are still unknown. Since long non-protein-coding RNAs (lncRNAs) play an important role in regulating multiple cellular functions, we used human endothelial cells treated with high glucose to mimic DM and with the uremic toxin indoxyl sulfate (IS) to mimic the endothelial injury associated with CKD. Differentially expressed lncRNAs in these conditions were analyzed by RNA sequencing. We discovered that lnc-SLC15A1-1 expression was significantly increased upon IS treatment in comparison with high glucose alone, and then cascaded the signal of chemokines CXCL10 and CXCL8 via sponging miR-27b, miR-297, and miR-150b. This novel pathway might be responsible for the endothelial inflammation implicated in augmenting CVD in DM/CKD and could be a therapeutic target with future clinical applications.

Semaphorin 5A suppresses the proliferation and migration of lung adenocarcinoma cells.

Semaphorin 5A (SEMA5A), a member of the semaphorin family, plays an important role in axonal guidance. Previously, the authors identified another possible role of SEMA5A as a prognostic biomarker for non­smoking women with lung adenocarcinoma in Taiwan, and this phenomenon has been validated in other ethnic groups. However, the functional significance of SEMA5A in lung adenocarcinoma remains unclear. Therefore, we assessed the function of SEMA5A in three lung adenocarcinoma cell lines in this study. Kaplan­Meier Plotter for lung cancer was conducted for survival analyses. Reverse transcription­quantitative PCR (RT­qPCR) and western blot analysis were performed to investigate the expression and post­translational regulation of SEMA5A in lung adenocarcinoma cell lines. A pre­designed PyroMark CpG assay and 5­aza­2'­deoxycytidine treatment were used to measure the methylation levels of SEMA5A. The biological functions of lung adenocarcinoma cells overexpressing SEMA5A were investigated by microarrays, and validated both in vitro (proliferation, colony formation and migration assays) and in vivo (tumor xenografts) experiments. The results revealed that the hypermethylation of SEMA5A and the cleavage of the extracellular domain of SEMA5A were responsible for the downregulation of the SEMA5A levels in lung adenocarcinoma cells (A549 and H1299) as compared to the normal controls. Functional analysis of SEMA5A­regulated genes revealed that they were involved in cellular growth and proliferation. The overexpression of SEMA5A in A549 and H1299 cells significantly decreased the proliferation (P<0.01), colony formation (P<0.001) and migratory ability (P<0.01) of the cells. The suppressive effects of SEMA5A on the proliferative and migratory ability of the cells were also observed in both in vitro and in vivo experiments using brain metastatic Bm7 lung adenocarcinoma cells. On the whole, the findings of this study suggest a suppressive role for SEMA5A in lung adenocarcinoma involving the inhibition of the proliferation and migration of lung transformed cells.

Identifying the functions and biomarkers of Codonopsis pilosula and Astragalus membranaceus aqueous extracts in hepatic cells.

BACKGROUND: Homeostasis is a crucial concept used to describe the condition of patients and the roles of herbs in traditional Chinese medicine. Qi-deficiency pattern is one of the conditions when loss of homeostasis and is usually characterized by symptoms including lassitude, spontaneous sweating, and a weak pulse, which are not easy to quantitate. Codonopsis pilosula and Astragalus membranaceus were usually prescribed for carriers with hepatitis and patients with metastatic colon cancer, because these patients tended to experience fatigue. However, crude drugs were prescribed based on the exterior symptoms of patients without controlling clinical setting, such as gender, age, and dietary habits. Limited molecular evidence of using gene expression as the guide for description is available. Therefore, the purpose of this study was to identify potential and objective biomarkers of these two qi-related drugs in a simplified cellular system. METHODS: Aqueous extracts of crude qi-tonifying herbs, C. pilosula and A. membranaceus, and that of a qi-consuming drug, Citrus reticulata, were prepared. Human liver cancer HepG2 cells were treated with the extracts of qi-tonifying herbs for 24 h. Differentially expressed genes were identified using microarrays and quantitative RT-PCR (qRT-PCR) and validated in two other hepatocellular cell lines, Huh7 and L-02. RESULTS: A total of 67 differentially expressed probes that responded to both herbs were identified. A pathway analysis revealed that these genes were involved in the development, growth, movement, and viability of the liver cells. CONCLUSIONS: After qRT-PCR validation and examination of clinical data from public domains, our results showed that two genes, GDF15 and HMOX1, could serve as biomarkers in liver cells for identifying responses after treatment with C. pilosula and A. membranaceus.

Macrophage Migration Inhibitory Factor Acts as the Potential Target of a Newly Synthesized Compound, 1-(9'-methyl-3'-carbazole)-3, 4-dihydro-ß-carboline.

For a newly synthesized compound, identifying its target protein is a slow but pivotal step toward understand its pharmacologic mechanism. In this study, we systemically synthesized novel manzamine derivatives and chose 1-(9'-methyl-3'-carbazole)-3, 4-dihydro-ß-carboline (MCDC) as an example to identify its target protein and function. MCDC had potent toxicity against several cancer cells. To identify its target protein, we first used a docking screen to predict macrophage migration inhibitory factor (MIF) as the potential target. Biochemical experiments, including mutation analysis and hydrogen-deuterium exchange assays, validated the binding of MCDC to MIF. Furthermore, MCDC was shown by microarrays to interfere with the cell cycle of breast cancer MCF7 cells. The activated signaling pathways included AKT phosphorylation and S phase-related proteins. Our results showed MIF as a potential direct target of a newly synthesized manzamine derivative, MCDC, and its pharmacologic mechanisms.

The Chinese medicine Kuan-Sin-Yin improves liver function in patients with chronic hepatitis C: A randomised and placebo-controlled trial.

BACKGROUND & AIMS: This study examined the effects of a traditional Chinese medicine decoction, Kuan-Sin-Yin (KSY), on patients with chronic hepatitis C (CHC) in a randomised and placebo-controlled clinical trial. METHODS: This trial enrolled 70 subjects with CHC who were randomised into 2 groups each with 35 participants. In total, 29 participants in the therapeutic group took 100mL of the herbal decoction daily, whereas 28 in the control group took an herbal placebo with the same dose and frequency for the 6-week study. The primary outcomes were liver function and viral load. Secondary measurements included haematopoietic and biochemical profiles, safety parameters, and a quality of life survey. All measurements were collected at the beginning of the study and after 6 weeks. RESULTS: In within-group analysis, significant decreases of glutamate pyruvate transaminase (GPT) 31.7±75.2IU/L and glutamate oxaloacetate transaminase (GOT) 20.3±45.7IU/L were found in the KSY group (p=0.031 and 0.024, respectively). In the between-group analysis, KSY reduced serum GOT and GPT levels by more than 20IU/L (p=0.027 and 0.047, respectively). KSY also significantly decreased viral load by 0.3 log units (p=0.047). In addition, KSY significantly decreased serum triglyceride 16.9±27.5mg/dL (p=0.024). CONCLUSIONS: This study demonstrates that taking the KSY herbal decoction for 6 weeks improves liver function and serum triglyceride levels and is safe for patients with CHC. The potential long-term effects of KSY on lipid metabolism related hepatoprotection and viral clearance warrant further investigation.

A Chinese Decoction, Kuan-Sin-Yin, Improves Autonomic Function and Cancer-Related Symptoms of Metastatic Colon Cancer.

BACKGROUND: Kuan-Sin-Yin (KSY) is a traditional Chinese medicine (TCM) decoction, which has been shown to have cytostatic effects on cancer cells and involved in the TCM theory of promoting yin-yang balance.Sonce many cancer patients suffer from autonomic dysfunction (AD), which correspond to yin-yang imbalance in TCM. The aim of this study is to evaluate the possible effect of KSY in metastatic colon cancer (mCRC) patients with AD. METHODS: We conducted a single-group experiment. Total 52 qualified patients were enrolled. Participants took the KSY daily for 2 weeks. The primary outcome was KSY efficacy as reflected in the heart rate variability (HRV) and electrical conductivity (µA) over 12 meridian points. Autonomic function was examined before and after the KSY intervention. The vagal and sympathetic tone were recorded by HRV; 12 meridian energies were measured using a meridian energy analysis device. Secondary outcomes were cancer-related symptoms and patient quality of life (QoL). RESULTS: The results showed that the KSY intervention improved AD via increasing the vagal tone (HF: P = .041), but not the sympathetic tone (LF: P = .154); total autonomic activity was significantly enhanced (HRV activity: P = .013). Intriguingly, energy increased more over the yin meridian (P = .010) than over the yang meridian (P = .015). Cancer-related symptoms and QoL were significantly improved (P < .05). CONCLUSION: The safety and effectiveness of KSY in improving AD in mCRC patients are through regulating the vagal-sympathetic dynamic balance, which correspond to the TCM yin-yang concept of energy.

Glucocorticoid protects hepatoma cells against metabolic stress-induced cell death.

Hepatocellular carcinoma is one of the most common cancers in the world. Previously, we found that the level of glucocorticoid receptor was significantly higher in hepatocellular carcinoma than in adjacent liver tissues. Moreover, in vitro and in vivo studies showed that glucocorticoid stimulated the growth of hepatoma cells. On the other hand, endogenous metabolites such as 2-methoxyestradiol, a metabolite of estrogen produced in liver, and lactic acid, an end-product of glycolysis can result in apoptosis of tumor cells. There are studies that glucocorticoid inhibited apoptosis induced by different chemotherapeutic drugs, whether glucocorticoid could block endogenous stresses, such as 2-methoxyestradiol- or lactic acid-induced apoptosis in human and murine hepatoma cells is not known. In this study, the antagonistic effects of dexamethasone on 2-methoxyestradiol- and lactic acid-induced apoptosis were investigated in human HepG2 and murine Hepa1-6 hepatoma cells. Treatment of hepatoma cells with 2.5-10 microM 2-methoxyestradiol or 25 mM lactic acid resulted in growth inhibition and decreased viability. In addition, results of cell cycle analysis, annexin V binding assay and DNA fragmentation formation showed that 2-methoxyestradiol- or lactic acid-induced apoptosis of hepatoma cells but these effects were partially blocked by dexamethasone. Combined treatment of hepatoma cells with dexamethasone and 2-methoxyestradiol or lactic acid partially reduced the 2-methoxyestradiol- or lactic acid-induced apoptosis signal. Treatment of hepatoma cells with 2-methoxyestradiol or lactic acid resulted in up-regulation of caspase-8, -9 and -3. Dexamethasone partially suppressed the caspase expression. The Bcl-2 level was induced by dexamethasone treatment but decreased after treatment with 2-methoxyestradiol or lactic acid. These results together suggest that glucocorticoids may protect hepatoma cells from metabolic stress-induced cell damage via anti-apoptotic pathways.