Lichen pectin-containing polysaccharide from Xanthoria elegans and its ability to effectively protect LX-2 cells from H2O2-induced oxidative damage.

Xanthoria elegans, a drought-tolerant lichen, is the original plant of the traditional Chinese medicine "Shihua" and effectively treats a variety of liver diseases. However, thus far, the hepatoprotective effects of polysaccharides, the most important chemical constituents of X. elegans, have not been determined. The aim of this study was to screen the polysaccharide fraction for hepatoprotective activity by using free radical scavenging assays and a H2O2-induced Lieming Xu-2 cell (LX-2) oxidative damage model and to elucidate the chemical composition of the bioactive polysaccharide fraction. In the present study, three polysaccharide fractions (XEP-50, XEP-70 and XEP-90) were obtained from X. elegans by hot-water extraction, DEAE-cellulose anion exchange chromatography separation and ethanol gradient precipitation. Among the three polysaccharide fractions, XEP-70 exhibited the best antioxidant activity in free radical scavenging capacity and reducing power assays. Structural studies showed that XEP-70 was a pectin-containing heteropolysaccharide fraction that was composed mainly of (1 â†’ 4)-linked and (1 â†’ 4,6)-linked α-D-Glcp, (1 â†’ 4)-linked α-D-GalpA, (1 â†’ 2)-linked, (1 â†’ 6)-linked and (1 â†’ 2,6)-linked α-D-Manp, and (1 â†’ 6)-linked and (1 â†’ 2,6)-linked ß-D-Galf. Furthermore, XEP-70 exhibited effectively protect LX-2 cells against H2O2-induced oxidative damage by enhancing cellular antioxidant capacity by activating the Nrf2/Keap1/ARE signaling pathway. Thus, XEP-70 has good potential to protect hepatic stellate cells against oxidative damage.

Hirsutella sinensis mycelium polysaccharides attenuate the TGF-ß1-induced epithelial-mesenchymal transition in human intrahepatic bile duct epithelial cells.

Hirsutella sinensis is the anamorph of Ophiocordyceps sinensis, and its mycelia has been used to effectively treat a variety of hepatobiliary diseases in clinical practice. In the present study, we performed a systematic study on the composition and structure of its polysaccharides, and then employed a TGF-ß1-induced human intrahepatic bile duct epithelial cell-epithelial-mesenchymal transition (HIBEC-EMT) model to investigate their effects on treating primary biliary cholangitis (PBC) based on hepatic bile duct fibrosis. Four polysaccharide fractions were obtained from H. sinensis mycelia by hot-water extraction, DEAE-cellulose column and gradient ethanol precipitation separation. HSWP-1a was an α-(1,4)-D-glucan; HSWP-1b and HSWP-1d mainly consisted of mannoglucans with a backbone composed of 1,4-linked α-D-Glcp and 1,4,6-linked α-D-Manp residues branched at O-6 of the 1,4-linked α-D-Glcp with a 1-linked α-D-Glcp as a side chain; and HSWP-1c mainly contained galactomannoglucans. These polysaccharide fractions protected HIBECs from a TGF-ß1-induced EMT, according to HIBEC morphological changes, cell viability, decreased E-cadherin and ZO-1 expression, and increased vimentin and collagen I expression. Furthermore, the effects of the polysaccharides might be mediated by inhibiting the activation of the TGF-ß/Smad signaling pathway, which attenuated hepatic bile duct fibrosis and potential PBC effects.

Tibetan mineral-herbal medicine Zuotai alleviates the depressive-like behaviors in chronic restraint-stressed mice while regulating stress hormone, inflammation and monoamine.

Introduction: Zuotai is an ancient mineral-herbal mixture containing ß-HgS in Tibetan medicine. It is used to treat nervous system diseases, similar to Chinese medicine cinnabar and Indian Ayurveda medicine Rasasindura. However, one of the key problems faced by Zuotai is that its indications are ambiguous. Our previous study found that Zuotai exhibited the activity of ameliorating depressive-like behaviors in a chronic mild stress model. However, due to the inherent limitations of animal models in simulating human disease, clear results often require more than one model for confirmation. Methods: Therefore, another depression model, chronic restraint stressed (CRS) mice, was used to validate the antidepression effect of Zuotai. Prophylactic treatment was conducted for 21 consecutive days while mice were subjected to chronic restraint stress. Results: It was observed that Zuotai and ß-HgS alleviated anhedonia, behavioral despair, stereotype behavior, and reduced exploratory and spontaneous movement in CRS mice. Zuotai and ß-HgS also reversed the increases of stress hormone corticosterone (Cort) in serum and pro-inflammatory cytokines in serum and brain, and increased the serotonin in cortex in CRS mice, with positive dose-effect relationship. The number of Ki67-positive cells in the dentate gyrus and the level of brain-derived neurotrophic factor (BDNF) in the hippocampus were slightly elevated in CRS mice treated with Zuotai; however, there was no statistically significant difference. Although Zuotai increased the total Hg concentration in main organs, the levels remained below those needed to result in observed adverse effect, at least for kidney and liver; and Zuotai showed no observed adverse effect on the brain histopathology, the cell proliferation in dentate gyrus, as well as the hippocampal and cortical organ coefficients. Conclusion: Zuotai exhibited the alleviation of depressive-like behaviors in CRS mice, accompanying with ameliorating stress hormone, peripherical and cerebral inflammation, and monoamine neurotransmitter.

Bioinformatics analysis and in vivo validation study of Ophiocordyceps sinensis (Berk.)G.H.Sungetal against lung adenocarcinoma.

ETHNOPHARMACOLOGICAL RELEVANCE: Lung adenocarcinoma (LUAD) is one of the main types of lung cancer. Ophiocordyceps sinensis has many potentially useful pharmacologic features, such as lung protection, and both anti-inflammatory and antioxidant activities. AIM OF THE STUDY: This study was conducted to investigate-using bioinformatics and in vivo experimental validation-the possible role of O. sinensis against LUAD. MATERIALS AND METHODS: We obtained important targets of O. sinensis for the treatment of LUAD using network pharmacology techniques and deep mining of the TCGA database, and validated them by molecular docking techniques and in vivo experiments. RESULTS: Through bioinformatics analysis and research, we screened BRCA1 and CCNE1 as important biomarkers for LUAD and as core targets of O. sinensis against LUAD. The non-small cell lung cancer signaling pathway, PI3K-Akt signaling pathway, and HIF-1 signaling pathway are potentially important pathways of O. sinensis against LUAD. The molecular docking results showed good binding between the active components in O. sinensis and the two core targets, and the in vivo experimental validation results indicated that O. sinensis had good inhibitory effects in the Lewis lung cancer (LLC) model. CONCLUSIONS: BRCA1 and CCNE1 are crucial biomarkers for LUAD and are important targets for O. sinensis to exert anti-LUAD effects.

The influence of genetic and acquired factors on the vulnerability to develop depression: a review.

Depression is one of the most common mental disorders that affects hundreds of millions of people worldwide and has claimed tens of thousands of lives. The causes are divided into two main areas: congenital genetic factors and acquired environmental factors. Congenital factors include genetic mutations and epigenetic events; acquired factors include birth patterns, feeding patterns, dietary patterns, childhood experiences, education and economic levels, isolation due to epidemics, and many other complex factors. According to studies, these factors play important roles in depression. Therefore, here, we analyze and study the factors from two aspects, describe their influence on individual depression, and analyze their underlying mechanisms. The results showed that both innate and acquired factors have significant effects on the occurrence of depressive disorder, and these findings may provide new ideas and methods for the study of depressive disorder, thus facilitating the process of depression prevention and treatment.

Effects of Tibetan medicine metacinnabar (ß-HgS) combined with imipramine or sertraline on depression-like symptoms in mice.

Depression is a common mood disorder that has exhibited an increased incidence rate worldwide, but the overall clinical efficacy of antidepressants remains unsatisfactory. In traditional Ayurveda and Tibetan medicines, ß-HgS-containing medicines have been used to treat neurological diseases for thousands of years, and our previous study found that ß-HgS ameliorated depression-like behaviors in chronic restraint stress (CRS)-treated or chronic unpredictable mild stress (CUMS)-treated mice. Hence, present study investigated the effects of ß-HgS combined with the clinical first-line antidepressants, imipramine (IMI) and sertraline (SER), on depression-like symptoms in CRS- and CUMS-co-treated mice. Our results revealed that ß-HgS promoted the antidepressant effect of SER on depression-like behavior in mice, and enhanced its effects on promoting glucocorticoid receptor (GR) expression and neuronal proliferation in key hippocampal subregions, as well as increasing interleukin 10 (IL-10) levels and decreasing malondialdehyde levels in the sera of stress-stimulated mice. As for IMI, ß-HgS enhanced its effects on preventing atrophy and severe structural damage in the hippocampus, as well as in promoting hippocampal GR levels and neuronal proliferation and serum IL-10 and superoxide dismutase (SOD) levels. Additionally, combination therapy resulted in the increased diversity of important intestinal microbiota compared to that of monotherapy, which may help sustain the health of the digestive tract and reduce inflammation to further enhance the antidepressant effects of IMI and SER in mice.

Exploring the mechanisms of action of Cordyceps sinensis for the treatment of depression using network pharmacology and molecular docking.

Background: Depression is the most common type of psychological disorder, with continuous, prolonged, and persistent bad moods as the main clinical feature. Cordyceps sinensis is a complex consisting of the ascospores and bodies of insect larvae from the Hepialidae family that have been parasitized by Cordyceps sinensis militaris. Previous studies have reported that this herb has antidepressant activity. The present study used network pharmacology and molecular docking techniques to investigate the potential antidepressant mechanisms of Cordyceps sinensis. Methods: The active ingredients of Cordyceps sinensis were identified using the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) and the potential targets were predicted using the PharmMapper platform. The GeneCards database was then used to obtain sub-targets for depression. Common targets were screened and enrichment analyses were performed using the Metascape platform. Finally, the relationship between the active ingredients and the core targets were verified by molecular docking. Results: Through network pharmacological analysis, 7 active ingredients in Cordyceps sinensis and 41 common targets of drugs and diseases were identified. The active ingredients of Cordyceps sinensis may exert antidepressant effects by acting on important targets such as catalase (CAT), CREB binding protein (CREBBP), epidermal growth factor (EGF), and E1A binding protein P300 (EP300), and by modulating the signaling pathways in which these targets are involved. Subsequently, the core targets were docked to the active ingredients and good binding was observed. Conclusions: The active ingredients of Cordyceps sinensis may exert antidepressant effects by regulating the CREB binding protein and anti-oxidative stress effects. The foxo signaling pathway (hsa04068), hypoxia-inducible factor 1 (HIF-1) signaling pathway (hsa04066), and Huntington's disease (hsa05016) may be involved in the underlying mechanisms of Cordyceps sinensis. The joint application of network pharmacology and molecular docking provides a new approach to study the mechanisms of action of traditional Chinese medicine. Cordyceps sinensis may play an important role in the future treatment of patients with depression.

Anti-Influenza Activity of Medicinal Material Extracts from Qinghai-Tibet Plateau.

To discover sources for novel anti-influenza drugs, we evaluated the antiviral potential of nine extracts from eight medicinal plants and one mushroom (Avena sativa L., Hordeum vulgare Linn. var. nudum Hook. f., Hippophae rhamnoides Linn., Lycium ruthenicum Murr., Nitraria tangutorum Bobr., Nitraria tangutorum Bobr. by-products, Potentilla anserina L., Cladina rangiferina (L.) Nyl., and Armillaria luteo-virens) from the Qinghai-Tibetan plateau against the influenza A/H3N2 virus. Concentrations lower than 125 µg/mL of all extracts demonstrated no significant toxicity in MDCK cells. During screening, seven extracts (A. sativa, H. vulgare, H. rhamnoides, L. ruthenicum, N. tangutorum, C. rangiferina, and A. luteo-virens) exhibited antiviral activity, especially the water-soluble polysaccharide from the fruit body of the mushroom A. luteo-virens. These extracts significantly reduced the infectivity of the human influenza A/H3N2 virus in vitro when used at concentrations of 15.6-125 µg/mL. Two extracts (N. tangutorum by-products and P. anserina) had no A/H3N2 virus inhibitory activity. Notably, the extract obtained from the fruits of N. tangutorum and N. tangutorum by-products exhibited different anti-influenza effects. The results suggest that extracts of A. sativa, H. vulgare, H. rhamnoides, L. ruthenicum, N. tangutorum, C. rangiferina, and A. luteo-virens contain substances with antiviral activity, and may be promising sources of new antiviral drugs.

Structure and immunomodulatory activity of a water-soluble α-glucan from Hirsutella sinensis mycelia.

Hirsutella sinensis, the anamorph of Ophiocordyceps sinensis (syn. Cordyceps sinensis), is a great substitute for precious and rare wild Cordyceps sinensis to effectively treat a variety of lung and kidney diseases. In this study, an α-glucan (named as HSWP-2a) was obtained by hot water extraction, DEAE-cellulose separation, and Sepharose CL-6B purification from H. sinensis mycelia. Different from known α-glucans, HSWP-2a is an α-(1 â†’ 4)-D-glucan that branched at O-6, O-3, or O-2 with a terminal 1-linked α-D-Glcp as side chain, with an average molecular weight of 870.70 kDa. Immunological tests showed that HSWP-2a could remarkably enhance the phagocytosis of macrophages and increase the production of NO, IL-1ß, IL-6, and TNF-α, via activating the p38, JNK, and NF-κB signaling pathways. Moreover, HSWP-2a could significantly promote splenic lymphocyte proliferation. Taken together, HSWP-2a may be potentially utilized as a natural immunomodulatory agent.

Bioactive Ingredients from Nitraria tangutorun Bobr. Protect Against Cerebral Ischemia/Reperfusion Injury Through Attenuation of Oxidative Stress and the Inflammatory Response.

Nitraria tangutorun Bobr. has been used for thousands of years as a native folk medicine to alleviate dizziness and neurasthenia due to oxygen. In our previous study, natural antioxidant components (namely, NJBE) were isolated from industrial N. tangutorun Bobr. juice byproducts (NJBE) from the Qinghai-Tibet plateau. The current investigation assessed the effects of NJBE on ischemic stroke in mice and the potential mechanisms. C57BL/6 mice received NJBE (25, 50, or 100 mg/Kg) by gavage for 14 days and then stroke was induced by the middle cerebral artery occlusion (MCAO) model, followed by reperfusion for 72 h. The evaluation of brain infarct size, behavioral tests, and functional assessments was conducted to assess the effects of NJBE after MCAO. Our results suggested that NJBE significantly decreases infarct size, improves neurological deficits, as well as reduces the number of GFAP+ and Iba-1+ cells after MCAO. NJBE inhibited nitric oxide and malondialdehyde production in the ischemic brain. Meanwhile, it attenuated the expressions of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx). Also, NJBE significantly attenuated the expression levels of proinflammatory indicators, including TNF-α, IL-1ß, IL-6, and IL-12. This process was accompanied by the downregulation of TLR4, TRAF6, pIκB/pIκB, and MMP9 expression and the upregulation of claudin-5 expression. NJBE induced improvements in brain injury. The neuroprotective effect of NJBE provides evidence for its potential application in stroke treatment.

Conjugated linoleic acid prompts bone formation in ovariectomized osteoporotic rats and weakens osteoclast formation after treatment with ultraviolet B.

BACKGROUND: Ultraviolet B (UVB) has been reported to prevent bone loss by promoting the synthesis of vitamin D. However, UVB can also enhance osteoclastic differentiation, inhibit osteogenic differentiation, and cause oxidative damage. The present study aimed to analyze the osteoprotective effects of UVB and conjugated linoleic acid (CLA) in rats with ovariectomy-induced osteoporosis, and to determine the interactions between UVB and CLA and their effects on bone mesenchymal stem cells (BMSCs) and bone marrow mononuclear cells (BMMCs). METHODS: In vitro, the distance of UVB irradiation and the dose of CLA were selected by immunofluorescence assays and Cytotoxicity assay. BMSCs and BMMCs were detected by immunohistochemical and immunofluorescence assays. In vivo, three-month-old female Sprague-Dawley rats that had undergone ovariectomy were treated with UVB and CLA. After 8 weeks of therapy, the femurs of the rats were examined by micro-computed tomography (CT) and immunohistochemical detection to assess the therapeutic efficacy. RESULTS: The least inhibitive UVB distance and dosage of CLA were selected for the in vivo experiments. CLA effectively weakened the osteogenic inhibitory effect of UVB (72 cm), significantly improved the activity of alkaline phosphatase (ALP), promoted the formation of mineralized nodules, and alleviated the oxidative damage induced by UVB. CLA also effectively weakened the osteoclast-promoting effect of UVB (72 cm), inhibited osteoclast formation, and inhibited the inflammatory damage to BMMCs caused by UVB (72 cm) irradiation. Micro-CT results showed that UVB irradiation could promote bone formation in ovariectomized Sprague-Dawley rats, while CLA could significantly promote bone regeneration. Immunofluorescence assays results showed that CLA alleviated UVB-induced oxidative damage to osteoblasts. The ROS detection results demonstrated that CLA effectively alleviated UVB-induced oxidative damage to BMSCs. Furthermore, Immunohistochemical assays showed that UVB and CLA treatment increased bone density, inhibited osteolytic osteolysis, and enhanced osteogenic activity. CONCLUSIONS: CLA can effectively weaken osteoclast promotion, osteogenic inhibition, and oxidative damage caused by UVB. Combination treatment of UVB and CLA exerts an osteoprotective effect on ovariectomized osteoporotic rats and stimulates osteogenesis. The molecular mechanism of this interaction requires further investigation.

Arsenic Content, Speciation, and Distribution in Wild Cordyceps sinensis.

The excessive arsenic content in wild Cordyceps sinensis has caused great concerns on human health. The toxicity of arsenic depends on its concentration, chemical form, and valence. The source studies of arsenic in C. sinensis are essential for safety evolution and quality control. We used ICP-MS and HPLC-ICP-MS methods to determine the total arsenic amount and the arsenic speciation. Synchrotron-based XANES and micro-XRF imaging techniques were used to characterize arsenic valence and distribution. The total arsenic amount range in wild C. sinensis samples was 5.77-13.20 µg/g with an average of 8.85 ± 2.5 µg/g. As(III) and As(V) were the main species in wild C. sinensis samples. The iAs only accounts for 4.47-11.42% of the extracted arsenic. Trivalent and pentavalent forms were the dominant chemical forms of arsenic. Besides, we found that arsenic was accumulated at the digestive tract of the host larva.

Neurotoxicity of ß-HgS differs from environmental mercury pollutants (MeHgCl and HgCl2) in Neuro-2a cell.

ß-HgS, differing from environmental mercury pollutants (MeHgCl and HgCl2) in chemical form, is used as traditional medicine in Asian countries for thousands of years. In this study, Neuro-2a cells were exposed to ß-HgS, MeHgCl and HgCl2 (5 µM) for 6-24 h. The cell viability of ß-HgS was higher than MeHgCl with 25.9% and 72.4% in 12 h and 24 h respectively. As the incubation time increased, MeHgCl had obvious damage to cell morphology, decreased the ratio of Bcl-2 and Bak and increased the expressions of TNF-α, IL-6 and IL-1ß significantly. Furthermore, the expressions of IL-1ß and IL-6 in HgCl2 group were increased significantly in 6 h and 24 h. The apoptotic rates in MeHgCl and HgCl2 group were respectively higher than ß-HgS with 32.2% and 7.30% in 24 h. Our findings indicate that ß-HgS is much less neurotoxicity than MeHgCl and HgCl2 in Neuro-2a cells.

Photocytotoxicity of white light-emitting diode irradiation on human lens epithelium and retinal pigment epithelium via the JNK and p38 MAPK signaling pathways.

WLEDs have lately been the preferred lighting device based on properties such as energy saving, high efficiency, longevity, and environmental protection. However, studies on the safety of white light-emitting diode (WLED) are limited. In our previous study, we found that WLED light (4000 K ± 500 K color temperature, 250 lx, and 20 min exposure) is photocytotoxic to three mammalian cell lines by causing cell lipid peroxidation. To further investigate the potential photocytotoxicity of WLEDs on the human body, we used two human eye cell lines SRA01/04 and D407 as target cells for evaluating its potential phototoxicity on the human eye in the present study based on cell viability, apoptosis, and intracellular oxidative stress assays, as well as the activation levels of reactive oxygen species (ROS)-related apoptosis pathways, including extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK) and p38 kinase (p38), using mitogen-activated protein kinase (MAPK) signaling pathway assays. The results showed that WLED light has photocytotoxicities on SRA01/04 and D407 cells, which were both in a time-, irradiance-, and color temperature-dependent manner and strongest at the conditions of 2 h irradiation time, 60 W/m2 irradiance, and 4000 K color temperature. Moreover, the photocytotoxicity of red light-emitting diode (LED) light was the strongest in the three tested monochromatic light compositions of WLED. Mechanism studies show that the potential phototoxicity of WLED on human lens epithelium and retinal pigment epithelium may be caused by its induced oxidative stress damage via the JNK and p38 MAPKs pathways.

Tobacco Mosaic Viral Nanoparticle Inhibited Osteoclastogenesis Through Inhibiting mTOR/AKT Signaling.

INTRODUCTION: Tobacco mosaic virus-based nanoparticles (TMV VNPs) were previously shown to promote osteogenic differentiation in vitro. This study aims to investigate whether and how TMV VNPs impact on osteoclastogenesis in vitro and bone injury healing in vivo. METHODS: Raw264.7 cells were cultured in osteoclastogenic medium in culture plates coated with or without TMV and TMV-RGD1 VNPs, followed by TRAP staining, RT-qPCR and WB assessing expression of osteoclastogenic marker genes, and immunofluorescence assessing NF-κB activation. TMV and TMV-RGD1-modified hyaluronic acid hydrogel were used to treat mouse tibial bone injury. Bone injury healing was checked by micro-CT and Masson staining. RESULTS: TMV and TMV-RGD1 VNPs significantly inhibited osteoclast differentiation and downregulated the expression of osteoclastogenic marker genes Ctr, Ctsk, Mmp-9, Rank, and Trap. Moreover, TMV and TMV-RGD1 VNPs inhibited NF-κB p65 phosphorylation and nuclear translocation, as well as activation of mTOR/AKT signaling pathway. TMV and TMV-RGD1-modified HA hydrogel strongly promoted mouse tibial bone injury with increased bone mass compared to plain HA hydrogel. The amount of osteoclasts was significantly reduced in TMV and TMV-RGD1 treated mice. TMV-RGD1 was more effective than TMV in inhibiting osteoclast differentiation and promoting bone injury repair. DISCUSSION: These data demonstrated the great potential of TMV VNPs to be developed into biomaterial for bone injury repair or replacement.

Ma-Nuo-Xi decoction has an immunostimulating effect in cyclophosphamide-immunosuppressed mice.

BACKGROUND: Ma-Nuo-Xi decoction (MNXD), as well as its hundreds of derivative preparations, has been used in Tibetan medicine since the 14th century. MNXD is in accordance with the theory of treatment determination based on syndrome differentiation. This study aimed to compare the effect of the auxiliary MNXD prescription (MNXD-AD) with that of the basic MNXD prescription (MNXD-BD) on the immunostimulating activity of MNXD. METHODS: The immunopotentiation of MNXD, MNXD-BD, and MNXD-AD was evaluated using a cyclophosphamide (CTX)-immunosuppressed mouse model. Their influences on non-specific and specific immunity were evaluated using immune organ indexes, peripheral white blood cell (WBC) count, red blood cell (RBC) count, platelet count, phagocytosis, macrophage-secreted nitric oxide (NO) and cytokines, natural killer (NK) cytotoxic activity, lymphocyte proliferation, serum cytokines, splenic T-lymphocyte subpopulations, and quantitative hemolysis of sheep red blood cell (QHS SRBC) assays. RESULTS: MNXD, MNXD-BD, and MNXD-AD increased the spleen and thymus indexes, as well as the peripheral WBC, RBC, and platelet counts. They also promoted phagocytosis, NO and cytokine secretion from macrophages, NK cytotoxic activity, and lymphocyte proliferation, and also raised the CD4+ /CD8+ T-cell ratio, serum cytokine concentrations, and haemolysin formation in CTX-treated immunosuppressed mice. Compared with MNXD-BD and MNXD-AD, MNXD was superior in restoring the phagocytic index, concanavalin A (ConA)-induced T-lymphocyte proliferation, NO secretion from macrophages, and haemolysin formation, as well as the levels of interleukin 1 beta (IL-1ß), and serum interleukin-2 (IL-2) and interferon gamma (INF-γ). CONCLUSIONS: MNXD, MNXD-BD, and MNXD-AD have excellent immunostimulating and myelosuppression-restoring activities on CTX-immunosuppressed mice. Among them, MNXD-AD might be an immunomodulator, which may happen to be in line with the clinical experience of Tibetan medicine physicians of using it to promote the efficacy of MNXD-BD.

Effect of combined chronic predictable and unpredictable stress on depression-like symptoms in mice.

BACKGROUND: Mental stress mainly induces depression, and predictable stress, as well as a constant bombardment of chronic unpredictable micro-stressors, always coexist in daily life. However, the combined effect of predictable and unpredictable stress on depression is still not fully understood. METHODS: The chronic restraint stress (CRS) is to restrain the mice for 6 h per day for 3 weeks, and the chronic unpredictable mild stress (CUMS) is to stimulate the mice with 7 different stressors for 3 weeks. We evaluated the combined effect of CRS and CUMS on depression-like symptoms using behavioral tests and investigated the action mechanism through analysis of neurotransmitters, brain-derived factors, inflammatory factors, antioxidants, and intestinal microorganisms. RESULTS: Our data suggested the combined stress of CRS and CUMS caused significant weight loss, food intake reduction, depression-like behaviors-including anhedonia, learned helplessness, and reduction in spontaneous activity-and even atrophy and severe structural damage to the hippocampus in mice. Our pathogenesis study showed that combined stress-induced the reduction of glucocorticoid receptor (GR) levels, loss of oligodendrocytes (NG2 and Olig2 cells), and inhibition of neuron proliferation in the CA1, CA3, and DG regions of the hippocampus, decreased the contents of monoamine neurotransmitters (5-HT and NE) and BDNF in the cerebral cortex, caused hyperactivity of the HPA system, led to immune dysfunction, aggravated oxidative stress, and weakened the capacity of antioxidants in mice. Compared with single stress, combined stress gave rise to a more significant diversity change of the gut microbiota. CONCLUSIONS: Combined stress caused significant depression-like behaviors, atrophy, and severe structural damage to the hippocampus in mice via monoamine neurotransmitter, BDNF, HPA axis, neurogenesis, and neurodegenerative, immune, oxidative stress and gut-brain axis action pathways.

Effects of ß-HgS on cell viability and intracellular oxidative stress in PC-12 cells.

Traditional Tibetan medicines containing ß-HgS have been used to treat chronic ailments for thousands of years. However, there has recently been speculation regarding the safety of these medicines due to their high mercury content. Although the toxic effect of ß-HgS has been previously investigated in vivo, the mechanism underlying the toxicity of this compound remains unclear. In this study, we investigate the mechanism of ß-HgS cytotoxicity via experiments performed on rat adrenal gland tumor cells (PC-12). Specifically, we analyze the viability and intracellular oxidative stress state of PC-12 cells treated with varying concentrations of ß-HgS. For comparison purposes, the effects of MeHgCl and HgCl2, two Hg-based compounds, on ROS generation and MDA, GSH/GSSG, Nrf2, NQO-1, and HO-1 levels are also determined. It should be noted that we used the small-molecule thiols of cell culture medium, such as cysteine, to increase the solubility of ß-HgS and prepare a ß-HgS solution to treat PC-12 cells. The obtained results show that ß-HgS inhibits cell viability at concentrations of 200-1000 ng Hg mL-1 (48 h treatment). In the concentration range of 200-600 ng Hg mL-1 (24 h treatment), the inhibitory effect of ß-HgS is stronger than that of MeHgCl; however, this trend is reversed at higher concentrations (800-1000 ng mL-1) and longer exposure times (48 h). Moreover, ß-HgS significantly promotes MDA, but has no appreciable influence on cell apoptosis and ROS generation in PC-12 cells, which suggests that its inhibitory effect on cell viability might be related to the stimulation of ROS-independent oxidative stress. Notably, ß-HgS and HgCl2 significantly increase the GSH content, GSH/GSSG ratio, NQO-1 mRNA expression, and HO-1 protein expression in PC-12 cells, indicating that the antioxidant protection against these compounds is triggered by Nrf2 activation. HPLC-AFS analysis shows that in ß-HgS and HgCl2 solutions, mercury exists in the same form of Hg2+, but the cytotoxicity of the former is greater. This is probably due to the additional oxidative damage induced by the S2- ion in ß-HgS. In conclusion, ß-HgS induces ROS-independent oxidative stress in PC-12 cells, and thus, is obviously cytotoxic. At the same time, it promotes the antioxidant capacity of cells by activating the Nrf2 pathway.

Immunostimulating effect of polysaccharides isolated from Ma-Nuo-Xi decoction in cyclophosphamide-immunosuppressed mice.

Ma-Nuo-Xi Decoction (MNXD) is well-known in Tibetan medicine to be in line with the theory of treatment determination based on syndrome differentiation. However, the components responsible for its immunomodulating effect are unknown. In this study, three polysaccharide components-MNXD-P, MNXD-BD-P, and MNXD-AD-P-were isolated from MNXD and its basic and auxiliary prescription decoctions, of which MNXD-BD-P is composed of ß-(1,4)-d-glucan and RG-I pectin, MNXD-AD-P contains mainly α-(1,4)-d-glucan and some amount of arabinogalactan and/or arabinorhamnogalactan, and MNXD-P contains components of both MNXD-BD-P and MNXD-AD-P. And treatment with these polysaccharides could significantly improve the host's specific and non-specific immunity, including cellular and humoral immunities, as well as promote recovery from myelosuppression in cyclophosphamide (CTX)-immunosuppressed mice. To our knowledge, this is the first report on chemical and immunoactivity study on polysaccharides from traditional Tibetan medicine compounds, which may provide a new idea for development of carbohydrate drugs from them.

Hormesis of methylmercury-human serum albumin conjugate on N9 microglia via ERK/MAPKs and STAT3 signaling pathways.

Methylmercury (MeHg+) is an extremely toxic organomercury cation that can induce severe neurological damage. Once it enters the body, methylmercury binds to amino acids or proteins containing free sulfhydryl groups. In particular, methylmercury is known to bind with human serum albumin (HSA) in human plasma; however, the effects of methylmercury-HSA conjugate (MeHg-HSA) on the central nervous system (CNS) are not fully understood. In the present study, we used the microglial cell line N9 as the target cells to evaluate the effect of MeHg-HSA on physiological function of the CNS preliminarily. The various factors in the cell culture were monitored by MTT assay, total lactate dehydrogenase assay, ELISA, qPCR, Western blot and flow cytometry techniques. The results showed that low-dose treatment with MeHg-HSA activated N9 cells, promoting cell proliferation and total cell number, enhancing NO and intracellular Ca2+ levels, and suppressing the release of TNFα and IL1ß without cytotoxic effects; while high-dose MeHg-HSA exhibited cytotoxic effects on N9 cells, including promoting cell death and increasing the secretion of TNFα and IL1ß. These results indicate that MeHg-HSA causes hormesis in microglia N9 cells. Furthermore, ERK/MAPKs and STAT3 signaling pathways related to the hormesis of MeHg-HSA on N9 cells. In addition, low dose of MeHg-HSA might be viewed as something very close to a lowest observed adverse effect level (LOAEL) for N9 cells. These findings will be useful for investigating the hormesis mechanism of MeHg+ and exploring the specific functions of MeHg-sulfhydryl conjugates on the central nervous system.