Synergistic Renoprotective Effects of Green Tea Extract and Gemigliptin against Tacrolimus-Induced Nephrotoxicity in Mice / Efectos Renoprotectores Sinérgicos del Extracto de Té Verde y Gemigliptina contra la Nefrotoxicidad Inducida por Tacrolimus en Ratones

SUMMARY: Although tacrolimus (TAC) significantly reduces allograft rejection incidence in solid-organ transplantation, its long-term use is associated with an increased risk of TAC-induced nephrotoxicity. In this study, we investigated the renoprotective effects of green tea extract (GTE) with or without the dipeptidyl peptidase 4 inhibitor, gemigliptin, by assessing serum creatinine levels, the amount of proteinuria, and histopathology in TAC-induced nephrotoxicity. TAC-induced nephrotoxicity was induced by intraperitoneal TAC injection, GTE was administered via subcutaneous injection, and gemigliptin was administered orally. Mice with TAC-induced nephrotoxicity exhibited a significant increase in both serum creatinine levels and 24-hour urine protein. However, when treated with GTE via subcutaneous injection, mice showed a decrease in serum creatinine levels and the amount of proteinuria. When GTE was combined with gemigliptin, further renoprotective effects were observed in biochemical assessments, consistent with the attenuation of TAC-induced nephrotoxicity in histopathology. The expression of p53 protein was lower in the mice treated with the combination of GTE and gemigliptin compared to mice with TAC-induced nephrotoxicity. Our results demonstrate that the combination of GTE and gemigliptin treatment reveals synergistic renoprotective effects by decreasing the expression of p53 protein. These findings suggest that the combination of GTE and gemigliptin could potentially be used as a prophylactic or therapeutic strategy for TAC-induced nephrotoxicity.

Aunque tacrolimus (TAC) reduce significativamente la incidencia de rechazo de aloinjertos en trasplantes de órganos sólidos, su uso a largo plazo se asocia con un mayor riesgo de nefrotoxicidad inducida por TAC. En este estudio, investigamos los efectos renoprotectores del extracto de té verde (GTE) con o sin el inhibidor de la dipeptidil peptidasa 4, gemigliptina, mediante la evaluación de los niveles de creatinina sérica, la cantidad de proteinuria y la histopatología en la nefrotoxicidad inducida por TAC. La nefrotoxicidad inducida por TAC se indujo mediante inyección intraperitoneal de TAC, el GTE se administró mediante inyección subcutánea y la gemigliptina se administró por vía oral. Los ratones con nefrotoxicidad inducida por TAC mostraron un aumento significativo tanto en los niveles de creatinina sérica como en la proteína en orina de 24 horas. Sin embargo, cuando se trataron con GTE mediante inyección subcutánea, los ratones mostraron una disminución en los niveles de creatinina sérica y en la cantidad de proteinuria. Cuando se combinó GTE con gemigliptina, se observaron efectos renoprotectores adicionales en las evaluaciones bioquímicas, lo que concuerda con la atenuación de la nefrotoxicidad inducida por TAC en histopatología. La expresión de la proteína p53 fue menor en los ratones tratados con la combinación de GTE y gemigliptina en comparación con los ratones con nefrotoxicidad inducida por TAC. Nuestros resultados demuestran que la combinación de tratamiento con GTE y gemigliptina revela efectos renoprotectores sinérgicos al disminuir la expresión de la proteína p53. Estos hallazgos sugieren que la combinación de GTE y gemigliptina podría usarse potencialmente como estrategia profiláctica o terapéutica para la nefrotoxicidad inducida por TAC.

Caragana sinica (Buc'hoz) Rehd. (jin ji er) polysaccharide regulates the immune function and intestinal microbiota of cyclophosphamide (CTX) induced immunosuppressed mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Caragana sinica (Buc'hoz) Rehd. is a plant widely grown in Yunnan, China, for both medicinal and edible purposes. The "National Compilation of Chinese Herbal Medicine" describes its nature as "slightly temperate and sweet". Caragana sinica is usually medicated with whole herbs, the main function is to replenish the kidneys and stop bleeding. Caragana sinica was used in folk medicine in Chuxiong, Yunnan, to treat deficiency colds, fatigue, fever, cough, hypertension, and other diseases. AIM OF THE STUDY: This article investigates the structural characteristics of Caragana sinica polysaccharide (CSP) and explores its immune-regulatory activity and molecular biological mechanisms in cyclophosphamide-induced immunosuppressed mice, as well as its effects on intestinal bacteria. METHODS: With the water-extraction and alcohol-precipitation method, Caragana sinica polysaccharide were extracted, obtaining CSP by purification. A variety of methods and techniques have been used to analyze the chemical properties and structural characteristics of CSP. Immunosuppressive mice model was established through intraperitoneal injection of cyclophosphamide (CTX) to study the immune-regulatory effects and mechanisms of CSP. RESULTS: The data indicated that CSP is a neutral heteropolysaccharide mainly composed of arabinose and galactose. This article uses immunosuppressive mice induced by cyclophosphamide (CTX) as the model. The results showed that CSP can promote the immune function of CTX treated immunosuppressed mice and regulate the diversity and composition of intestinal microbiota. CSP can increase macrophage phagocytosis, NK cell killing activity, and lymphocyte proliferation activity. It can also repair the index and morphological damage of the thymus and spleen. And by binding to the TLR4 receptor, MyD88 was activated and interacted with TRAF6 to promote the transfer of NF-κB into the nucleus. Thereby promoting cytokine release and increasing the production of IL-1ß, IL-6, IL-10, TNF-α, IgA, and IgG in the serum. CSP also effectively alleviated the liver damage caused by CTX through antioxidant activity. Furthermore, CSP can dramatically affect the intestinal microbiota and the body's immunity by boosting the relative presence of Bacteroides and Verrucamicrobiota. CONCLUSIONS: Research results indicated that CSP can regulate the immune function of mice, providing a basis for developing CSP as a potential immune modulator and functional food.

Study on alleviate effect of Wuzhi capsule (Schisandra sphenanthera Rehder & E.H. Wilson extract) against mycophenolate mofetil-induced intestinal injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Schisandra sphenanthera Rehder & E.H. Wilson (S. sphenanthera) is a botanical medicine included in the 2020 edition of the ChP that has a variety of medicinal activities, including hepatoprotective, anticancer, antioxidant and anti-inflammatory properties. Wuzhi capsule (WZ) is a proprietary Chinese medicine made from an ethanolic extract of S. sphenanthera that is commonly used to treat drug-induced liver injury. However, there are no research reports exploring the effects of WZ on the prevention of mycophenolate mofetil (MMF)-induced intestinal injury and its underlying mechanisms. AIM OF THE STUDY: This experiment aimed to evaluate the ameliorative effect of WZ on MMF-induced intestinal injury in mice and its underlying mechanisms. MATERIALS AND METHODS: A mouse model of MMF-induced intestinal injury was established and treated with WZ during the 21-day experimental period. The pathological characteristics of the mouse ileum were observed. Tight junction (TJ) protein changes were observed after immunofluorescence staining and transmission electron microscopy, and ROS levels were measured by using DHE fluorescent dye and the TUNEL assay for apoptosis. The expression of p65, p-p65, IκBα, p-IκBα, the TJ proteins occludin and ZO-1 and the apoptosis-related proteins Bax, Bcl-2, cleaved caspase-3 and caspase-3 were analysed by Western blot. Levels of DAO, ET, TNF-α, IL-1ß, IL-6, IFN-γ, MDA and SOD were analysed by using kits. RESULTS: MMF activated the NF-κB signaling pathway to cause intestinal inflammation, increased intestinal permeability, changed the expression of TJ protein in the intestinal epithelium, and increased oxidative stress and apoptosis levels. WZ significantly downregulated the expression of p-p65 and p-IκBα to relieve the inflammatory response, reduced intestinal permeability, maintained intestinal TJ protein expression, and reduced intestinal oxidative stress and apoptosis. CONCLUSION: Our research suggested that MMF can cause intestinal injury; by contrast, WZ may exert anti-inflammatory, antioxidant and apoptosis-reducing effects to alleviate MMF-induced intestinal injury.

The immune-enhancing effects of a mixture of Astragalus membranaceus (Fisch.) Bunge, Angelica gigas Nakai, and Trichosanthes Kirilowii (Maxim.) or its active constituent nodakenin.

ETHNOPHARMACOLOGICAL RELEVANCE: A mixture (SH003) of Astragalus membranaceus (Fisch.) Bunge, Angelica gigas Nakai, and Trichosanthes Kirilowii (Maxim.) has beneficial effects against several carcinomas. There have been few reports on an immune-enhancing activity of SH003 and its active constituent nodakenin. AIM OF THE STUDY: This study aimed at identifying the immune-enhancing effect of SH003 and nodakenin. MATERIALS AND METHODS: The immune-enhancing effect was evaluated using RAW264.7 macrophages, mouse primary splenocytes, and a cyclophosphamide (CP)-induced immunosuppression murine model. RESULTS: The results show that SH003 or nodakenin stimulated the production levels of granulocyte colony-stimulating factor, IL-12, IL-2, IL-6, TNF-α, and nitric oxide (NO) and the expression levels of iNOS in RAW264.7 macrophages. SH003 or nodakenin also enhanced NF-κB p65 activation in RAW264.7 macrophages. SH003 or nodakenin stimulated the production levels of IFN-γ, IL-12, IL-2, TNF-α, and NO and the expression levels of iNOS in splenocytes. SH003 or nodakenin increased the splenic lymphocyte proliferation and splenic NK cell activity. In addition, SH003 or nodakenin increased the levels of IFN-γ, IL-12, IL-2, IL-6, and TNF-α in the serum and spleen of CP-treated mice, alleviating CP-induced immunosuppression. CONCLUSION: Taken together, the results of this study show that SH003 improved immunosuppression through the activation of macrophages, splenocytes, and NK cells. These findings suggest that SH003 could be applied as a potential immunostimulatory agent for a variety of diseases caused or exacerbated by immunodeficiency.

Protective effects of selenium in tacrolimus-induced lung toxicity: potential role of heme oxygenase 1.

The present study aimed to evaluate the protective effects of selenium (Sel) administration against tacrolimus (Tac) - induced lung toxicity and to assess the relation between heme oxygenase 1 (HO-1) and these effects. The study was conducted on 36 Wistar male albino rats equally divided into four groups: (i) normal control; (ii) Sel (0.1 mg/kg per day p.o. for four weeks); (iii) TAC 3 mg/mL as single oral dose on 27th day; and (iv) Tac + Sel. Lung tissues, lung homogenate, and bronchoalveolar lavage of the sacrificed animals were investigated biochemically and histopathologically, by immunohistochemistry or by PCR. The Tac group showed significantly lower expression of HO-1. Administration of Sel was associated with increased HO-1 expression. Oxidative (malondialdehyde, reduced glutathione, superoxide dismutase, myeloperoxidase, and glutathione peroxidase activity) and nitrosative stress (nitric oxide) markers and markers of inflammation (interleukin 1ß (IL-1ß), IL-6, and IL-10) showed changes corresponding to HO-1 levels in rat groups. Tac group showed the highest expression of caspase-3. Sel exerted a protective role against Tac-induced lung toxicity.

Differences in Intestinal Metabolism of Ginseng Between Normal and Immunosuppressed Rats.

BACKGROUND AND OBJECTIVE: Ginseng is usually consumed as a dietary supplement for health care in the normal state or prescribed as a herbal medicine in pathologic conditions. Although metabolic studies of ginseng are commonly performed on healthy organisms, the metabolic characteristics in pathologic organisms remain unexplored. This study aimed to uncover the difference in intestinal metabolism of ginseng between normal and cyclophosphamide-induced immunosuppressed rats and further discuss the potential mechanisms involved. METHODS: Twelve Sprague-Dawley rats (6-8 weeks old) were randomly divided into two groups: the normal group (NG) and immunosuppressed group (ISG). Rats in the NG and ISG groups were intraperitoneally administered normal saline and cyclophosphamide injections (40 mg/kg) on the 1st, 2nd, 3rd and 10th days; on the 12th day, all rats were intragastrically administered ginseng water extract (900 mg/kg). The difference in intestinal metabolism of ginseng was compared using an ultra-high-performance liquid chromatography coupled with quadruple time-of-flight mass spectrometry-based metabolomics approach, and the diversities of gut microbiota were analyzed by 16S rRNA gene sequencing between the two groups. RESULTS: The intestinal metabolomic characteristics of ginseng were significantly different between the normal and immunosuppressed rats, with the ginsenoside F2 (F2), 20S-ginsenoside Rg3 (20(S)-Rg3), pseudo-ginsenoside Rt5 (Pseudo-Rt5), ginsenoside Rd (Rd), ginsenoside Rh1 (Rh1), 20S-ginsenoside Rg1 (20(S)-Rg1), ginsenoside compound K (CK), ginsenoside Rg2 (Rg2) and 20S-panaxatriol (S-PPT) more abundant in immunosuppressed ones (P < 0.05). Additionally, the composition of gut microbiota was remarkably altered in the two groups, with some specific bacterial communities such as Bacteroides spp., Eubacterium spp. and Lachnospiraceae_UCG-010 spp. increased and Bifidobacterium spp. decreased in immunosuppressed rats compared with normal ones. CONCLUSION: The intestinal metabolism of ginseng in immunosuppressed rats was significantly different from that in normal ones, which might be partly attributed to the changes in the intensity of specific gut bacteria. The outcomes of this study could provide scientific data for rationalization of ginseng use as both a dietary supplement and herbal medicine.

The effects of aqueous extract of Maca on energy metabolism and immunoregulation.

BACKGROUND: In the present work, we investigated the effects of aqueous extract of Maca (AEM) on energy metabolism and immunoregulation in spleen-deficient mice. METHOD: We established a cyclophosphamide-induced spleen-deficiency model with ginseng, a herb that strengthens splenic function, as a control. Sixty male Kunming mice were randomly divided among 5 groups: normal, model, ginseng control (1.5 g/kg), AEM high dose (1.5 g/kg), and AEM low dose (0.75 g/kg). All animals, except those in the normal group, were injected with cyclophosphamide to induce spleen deficiency. Furthermore, we investigated differences in the thermotropic behaviors of mice using the Animal Thermotropism Behavior Surveillance System to detect energy metabolism-related assays and immune regulation assays. RESULTS: Mice given AEM exhibited tropism in response to hot plate exposure. AEM inhibited loss of body weight and immune organ atrophy caused by cyclophosphamide, increased the cAMP/cGMP ratio in blood, and enhanced the activities of Na+-K+-ATPase, Ca2+-Mg2+-ATPase, lactate dehydrogenase, and hepatic glycogen. AEM significantly reversed declining white blood cells and platelet counts, and increased the hemoglobin content within peripheral blood cells. AEM improved the protein levels of IFN-γ, TNF-ß, IL-2, and IL-4 in the spleen. CONCLUSIONS: Maca possesses the Traditional Chinese Medicine (TCM) property of warm and appears to strengthen spleen function.

Protective effect of Nasturtium officinale R. Br and quercetin against cyclophosphamide-induced hepatotoxicity in rats.

Cyclophosphamide (CPA) is used in the management of autoimmune conditions and malignant illnesses. However, its therapeutic use is limited because of its severe side effects, especially hepatotoxicity attributed to oxidative stress. Nasturtium officinale R. Br (watercress or WC) has pharmacological properties, such as anti-inflammation, and antioxidant activities. Therefore, the present study was design to assess effects of WC or its active ingredient, quercetin (QE), against CPA-induced hepatotoxicity. For this study, 49 male Wistar rats (200-250 g) were randomly selected and categorized into seven equal groups. The animals were pre- and post-treated with both hydroalcoholic extract of WC (500 mg/kg) and quercetin (75 mg/kg) for 10 consecutive days, and intraperitoneal administration of CPA (200 mg/kg) was performed on only day 10, one hour before the last dose of WC or quercetin. On day 11, all the animals were sacrificed, and their blood and liver were gathered for evaluation of the liver enzyme, hepatic oxidative stress markers, antioxidant enzymes activity, and hematoxylin and eosin staining. CPA significantly increased malondialdehyde (MDA), protein carbonyl (PCO) and nitric oxide (NO) levels and liver biomarkers. Otherwise, hepatic catalase (CAT), reduced glutathione (GSH), total thiol content (tSH), and ferric reducing antioxidant power (FRAP) were considerably lower than the control group. Results showed that WC has the ability to reduce the changes (MDA, PCO, FRAP, CAT, ALT and AST) and QE (MDA, PCO, AST) induced by CPA (p < 0.05). Histopathological finding confirmed the indicated results. These findings propose that WC and QE have protective effect against the CPA-induced hepatotoxicity by decreasing oxidative stress.

The hepatoprotective effects of XCHD and MgIG against methotrexate-induced liver injury and inflammation in rats through suppressing the activation of AIM2 inflammasomes.

BACKGROUND: Recent studies have shown that drug-induced liver injury may be related to the immune response activated by drugs. A cytosolic dsDNA inflammasome called absent in melanoma 2 (AIM2) was found to be associated with aseptic inflammation. The present study aimed to explore the effects of on the liver injury and inflammation in methotrexate (Mtx)-induced rats. METHODS: Sprague Dawley (SD) rats were selected and classified into 4 groups randomly, includes control group, Mtx group, Mtx-Xiaochaihu decoction (XCHD) group and Mtx-magnesium isoglycyrrhizinate (MgIG) group. Light microscopy was used to examine histological specimens after hematoxylin-eosin (HE) staining. The AST levels in liver tissue and blood serum ALT in the rats were assessed with enzyme linked immunosorbent assay (ELISA). Then AIM2 expression and inflammatory factors, including caspase-1, IL-18, and IL-1ß, in the liver biopsy specimens of rats were detected by immunohistochemistry. Furthermore, the correlation between inflammatory and AIM2 expression factors was comprehensively analyzed. RESULTS: Functional and structural hepatotoxicity can be caused by the exposure to Mtx, which was supported by the improved biochemical marker levels and the worse histopathological changes in liver tissue. Compared with the Mtx group, the levels of liver enzymes ALT and AST, histological deterioration in the liver tissues were effectively decreased by XCHD and MgIG treatment, respectively. In addition, the expression of AIM2, caspase-1 and IL-1ß was observably higher in the Mtx group, which was apparently inhibited in the Mtx-XCHD and Mtx-MgIG groups. There was no obvious change in IL-18 expression among four groups. AIM2 expression were positively associated with the severity of liver inflammation and had a higher relevance with caspase-1 expression. CONCLUSIONS: AIM2 inflammasome in hepatocytes has a significant effect on the development of Mtx-induced liver injury, which can be ameliorated by both XCHD and MgIG treatment. The latent mechanism and potential signal pathway require further study.

Protective effect of Vitamin D3 against lead induced hepatotoxicity, oxidative stress, immunosuppressive and calcium homeostasis disorders in rat.

Lead (Pb) is an extremely poisonous, non-essential trace element and toxicity develops in humans following frequent exposure to the heavy metal in polluted environmental and occupational settings. Pb induces hepatic damage through the depletion of the antioxidant system, enhancing cellular oxidative stress and stimulation of proinflammatory cytokines. Although the antioxidant and anti-inflammatory actions of vitamin D3 (VD3) are well-established, a minority of studies measured the protective actions of VD3 against Pb toxicity. Therefore, this work studied the effects of vitamin VD3 therapy on the fundamental molecular basis underlying hepatic injury induced by chronic Pb toxicity. Twenty-four adult male rats were distributed equally into the negative controls (NC), positive controls (PC) and VD3 groups. While both the PC and VD3 groups received Pb-acetate in drinking water (1000 mg/L) for four weeks, the latter group also received intramuscular VD3 injections (1000 IU/kg; 3 days/week) simultaneously with Pb. The liver enzymes together with the serum and hepatic tissue Pb concentrations increased markedly in the PC group compared with the NC group. Pb toxicity also drastically induced hepatocyte apoptosis/necrosis, increased the hepatic tissue concentrations of malondialdehyde and the pro-inflammatory cytokines (TGF-ß, IL-4 & TNF-α) as well as reduced the anti-oxidative enzymes (GSH, GPx & CAT) and the anti-inflammatory cytokine, IL-10, compared with the NC group. Pb also significantly decreased the serum concentrations of VD3 and Ca2+. Additionally, the hepatic expressions of VD receptor, Cyp24a1 enzyme, L-type Ca2+-channel, calbindin-D28k & -D29k, calmodulin and calmodulin-dependent protein kinase II were significantly upregulated, whereas the VD binding protein, CYP2R1 enzyme and T-type Ca2+-channel were markedly inhibited at the gene and protein levels following Pb intoxication. VD3 alleviated the hepatic damage, inhibited the oxidative stress and pro-inflammatory molecules as well as upregulated the anti-oxidant and anti-inflammatory markers and restored the expression of the VD/Ca2+ regulatory molecules compared with the PC group. VD3 supplementation discloses promising protective effects against Pb-induced hepatic damage, through its anti-inflammatory and antioxidant actions as well as by modulating the hepatocyte calcium homeostatic molecules.

High-content analysis boosts identification of the initial cause of triptolide-induced hepatotoxicity.

Triptolide (TP) has been widely used in China for more than 40 years as an immunosuppressive agent. Recently, serious concerns have been raised over TP-induced liver injury, though the real hepatotoxic mechanism is still unclear, particularly in terms of the initial cause. To our knowledge, this study is the first to screen systematically the mechanism of TP-induced toxicity through a global cytotoxicity profile high-content analysis using three independent cytotoxic assay panels with multiple endpoints of cytotoxicity, including cell loss, mitochondrial membrane potential, nuclear membrane permeability, manganese superoxide dismutase, phosphorylated gamma-H2AX, light chain 3B, lysosome, reactive oxygen species and glutathione. We assessed nine parameters and four stress response pathway models by labeling nuclear factor erythroid 2-related factor 2, activating transcription factor 6, hypoxia inducible factor 1α and nuclear factor κB and found that all testing parameters except glutathione and manganese superoxide dismutase showed concentration- and time-dependent changes, as well as increased cell loss after TP treatment. Considering that RNA polymerase II is the molecular target of TP, we quantified transcription from inducible genes, bromodeoxyuridine incorporation, and expression from transiently transfected green fluorescence protein plasmids in HepG2 cells. The results show that inhibition of global transcription by TP took place at earlier times and at lower concentrations than those observed for cell death. Therefore, global transcriptional suppression and the cell dysfunction it drives play a central role in TP-induced hepatotoxicity. This provides valuable information for the safe use of TP in the clinic.

Baicalein ameliorates pristane-induced lupus nephritis via activating Nrf2/HO-1 in myeloid-derived suppressor cells.

INTRODUCTION: Lupus nephritis (LN) is a representative manifestation in systemic lupus erythematosus (SLE). Some studies have shown that myeloid-derived suppressor cells (MDSCs) play a vital role in the regulation of the SLE process. MDSC infiltration in the kidney as well as inflammation and oxidative stress provokes the acceleration and deterioration of LN. Nuclear factor E2-related factor 2 (Nrf2) is thought to be a major regulator of the antioxidant response. Baicalein is a flavonoid with known anti-inflammatory effects and antioxidant response. However, the effects of baicalein on MDSCs, inflammation, and oxidative stress are not evaluated in the development of pristane-induced LN in mice. METHODS: The renoprotective effect of baicalein was detected in a pristane-induced lupus mice model. NLRP3 inflammasome activation and NF-κB phosphorylation as well as reactive oxygen species (ROS) production and Nrf2 activation were examined. The percentages and function changes of MDSCs were measured. The possible mechanisms of the underlying effects of baicalein on ROS production and signaling pathways of Nrf2/heme-oxygenase (HO)-1, NLRP3 inflammasome, and NF-κB phosphorylation in lipopolysaccharide (LPS)-primed MDSCs were analyzed. RESULTS: Baicalein reduced proteinuria and attenuated renal function impairment and renal histopathology including intrinsic cell proliferation, cellular crescents, and podocyte injury as well as glomerulonephritis activity in lupus mice. Moreover, baicalein downregulated the activation of NLRP3 inflammasome and levels of ROS or NF-κB phosphorylation, and it enhanced Nrf2 activation. Of note, baicalein inhibited the expansion of MDSCs and improved the function of MDSCs in lupus mice. Through analyzing LPS-primed MDSCs in vitro, baicalein was found to exhibit cytoprotective effects coincident with the induction of Nrf2/HO-1 signaling and the suppression of the NLRP3 inflammasome. CONCLUSION: The data show that baicalein alleviates the symptoms of pristane-induced LN and suggest that the alleviation may be attributed to inhibition of MDSC expansion and regulation of the balance of the Nrf2/HO-1 signal and NLRP3 expression in MDSCs.

A Comparative Study on the Effects of Different Parts of Panax ginseng on the Immune Activity of Cyclophosphamide-Induced Immunosuppressed Mice.

The objective of the present study was to compare the effects of the immunological activity of various parts (root/stem/leaf/flower/seed) of five-year-old ginseng on the immune system of immunosuppressive mice. Immunosuppression was induced by cyclophosphamide (CTX) in the mouse model, whereas levamisole hydrochloride tablet (LTH) was used for the positive control group. We found that ginseng root (GRT), ginseng leaf (GLF), and ginseng flower (GFR) could relieve immunosuppression by increased viability of NK cells, enhanced immune organ index, improved cell-mediated immune response, increased content of CD4⁺ and ratio of CD4⁺/CD8⁺, and recovery of macrophage function, including carbon clearance, phagocytic rate, and phagocytic index, in immunodeficient mice. However, ginseng stem (GSM) and ginseng seed (GSD) could only enhance the thymus indices, carbon clearance, splenocyte proliferation, NK cell activities, and the level of IL-4 in immunosuppressed mice. In CTX-injected mice, GRT and GFR remarkably increased the protein expression of Nrf2, HO-1, NQO1, SOD1, SOD2, and CAT in the spleen. As expected, oral administration of GRT and GFR markedly enhanced the production of cytokines, such as IL-1ß, IL-4, IL-6, IFN-γ, and TNF-α, compared with the CTX-induced immunosuppressed mice, and GRT and GFR did this relatively better than GSM, GLF, and GSD. This study provides a theoretical basis for further study on different parts of ginseng.

The Immuno-Enhancement Effects of Tubiechong (Eupolyphaga sinensis) Lyophilized Powder in Cyclophosphamide-Induced Immunosuppressed Mice.

Tubiechong (Eupolyphaga sinensis) is an important material used in traditional Chinese medicine (TCM). However, the immunoregulation effects of E. sinensis Lyophilized Powder (ESL) are unclear. The in vivo study thus designed to elucidate the immuno-enhancement effects of ESL in immunosuppressed mice induced by cyclophosphamide (CTX). Mice were treated with three doses of ESL (0.5, 1.0 and 2.0 g/kg). Compared with model group, ESL notably increased the immune organ index, mononuclear macrophages function and the level of nature killer cell (NK) (p < 0.05 or p < 0.01), delayed type hypersensitivity (DTH) was also improved (p < 0.05). The level of superoxide dismutase (SOD) and catalase (CAT) were enhanced (p < 0.05), while malonyldialdehyde (MDA) and nitrogen monoxide (NO) were reduced (p < 0.05 or p < 0.01). Meanwhile, cluster determinant (CD)3+ T cell, CD4+ T cell and CD4+/CD8+ ratio were increased (p < 0.01). The cytokines secretion such as interleukin (IL)-2 and tumor necrosis factor alpha (TNF-α) were notably increased (p < 0.05 or p < 0.01), and IL-6 and IL-16 were also enhanced (p < 0.05). Furthermore, ESL significantly inhibited the phosphorylation of c-Jun N-terminal kinase (JNK), down-regulated the expression of Bcl-2 associated X protein (Bax), up-regulated the B cell lymphoma-2 protein (Bcl-2) expression and decreased the Bax/Bcl-2 ratio in spleen tissues (p < 0.05). In brief, all these findings suggest that ESL could effectively improve immune functions via modulating oxidative systems and innate immune cells. Abbreviations: TCM: Traditional Chinese Medicine; ESL: Eupolyphaga sinensis Lyophilized Powder; CCl4: Carbon tetrachloride; ERK: Extracellular regulated protein kinases; CTX: Cyclophosphamide; DTH: Delayed type hypersensitivity; SOD: Superoxide dismutase; CAT: Catalase; MDA: Malonyldialdehyde; NO: Nitrogen monoxide; NK: Nature killer cell; CD: Cluster determinant interleukin; TNF-α: Tumor Necrosis Factor alpha; JNK: c-Jun N-terminal kinase; Bax: Bcl-2 associated X protein; Bcl-2: B cell lymphoma-2 protein; Th1: Type-1 helper; Th2: Type-2 helper; FAMEs: Fatty acid methyl esters; DNFB: 2,4 - Dinitrofluorobenzene; ELISA: Enzyme-linked immuno sorbent assay; MAPK: Mitogen activated protein kinase; Cyt-c: Cytochrome c; SCFAs: Short-chain fatty acids; SDS-PAGE: Sodium dodecyl sulfate polyacrylamide gel electrophoresis.

Structure-immunomodulatory activity relationships of Hedysarum polysaccharides extracted by a method involving a complex enzyme combined with ultrasonication.

A new, more effective and environmentally friendly method involving a complex enzyme combined with ultrasonication was employed to extract and isolate three novel polysaccharides (HPS-MCs: HPS-MC, HPS-MC (50%) and HPS-MC (80%)) of Radix Hedysari. Compared with polysaccharides obtained using a traditional extraction method (hot water extraction, HPS-R), the yields and total carbohydrate contents of HPS-MCs were significantly higher. HPS-MC (80%) exhibited relatively strong immunomodulatory activity and a concentration-dependent dose-response relationship under cyclophosphamide (CP)-induced immunosuppressive conditions in mice models. To more comprehensively investigate the relationships between structural characteristics and immunomodulatory activity, HPS-MC (80%) was fractionated into three major homogeneous polysaccharide fractions (HPS-MC (80%)s: HPS-MC (80%)-1, HPS-MC (80%)-2, and HPS-MC (80%)-3). These three homogeneous polysaccharides had different mass percentages of monosaccharides species (rhamnose, arabinose, mannose, glucose, and galactose) by gas chromatography (GC) and different molecular weights and chain conformations by high-performance gel permeation chromatography coupled with multi-angle laser light scattering (HPGPC-MALLS), and promoted macrophage and splenocyte proliferation to different degrees. These findings indicated that HPS-MC (80%) had a prominent potential immune response, especially HPS-MC (80%)-2 and HPS-MC (80%)-3, and might be suitable candidates for functional foods or potential novel immunomodulators.

Olea europaea leaf extract up-regulates Nrf2/ARE/HO-1 signaling and attenuates cyclophosphamide-induced oxidative stress, inflammation and apoptosis in rat kidney.

Olive leaf extract (OLE) has potential health benefits and protects against cytotoxicity in different organs. However, nothing has yet been reported on its potential to prevent cyclophosphamide (CP)-induced nephrotoxicity. This study investigated the possible protective effect of OLE on CP-induced kidney injury in rats, focusing on oxidative stress, inflammation, apoptosis and Nrf2/ARE/HO-1 signaling. Rats received 100 or 200 mg/kg body weight OLE for 15 days and a single injection of 150 mg/kg CP at day 16. CP induced kidney injury evidenced by the significantly increased serum creatinine and urea, and histopathological alterations, including glomerular atrophy, interstitial hemorrhage, dilated urinary space and necrosis. CP-induced rats exhibited increased kidney lipid peroxidation, protein carbonyl, nitric oxide (NO) and pro-inflammatory cytokines, and up-regulated NF-κB, Bax, cytochrome c and caspase-3. OLE ameliorated kidney function markers and prevented CP-induced tissue damage. In addition, OLE significantly prevented oxidative stress, inflammation and apoptosis by enhancing the antioxidant defenses and Bcl-2 expression, and suppressing the pro-inflammatory and pro-apoptotic markers NF-κB, Bax, cytochrome c and caspase-3. OLE up-regulated Nrf2, HO-1 and NQO-1 expression in the kidney of CP-induced rats. In conclusion, OLE has a substantial protective role against CP-induced nephrotoxicity in rats by up-regulating the Nrf2/ARE/HO-1 signaling, enhancing the antioxidant activity and attenuating inflammation and apoptosis.

Investigation of the effect of hepatic metabolism on off-target cardiotoxicity in a multi-organ human-on-a-chip system.

Regulation of cosmetic testing and poor predictivity of preclinical drug studies has spurred efforts to develop new methods for systemic toxicity. Current in vitro assays do not fully represent physiology, often lacking xenobiotic metabolism. Functional human multi-organ systems containing iPSC derived cardiomyocytes and primary hepatocytes were maintained under flow using a low-volume pumpless system in a serum-free medium. The functional readouts for contractile force and electrical conductivity enabled the non-invasive study of cardiac function. The presence of the hepatocytes in the system induced cardiotoxic effects from cyclophosphamide and reduced them for terfenadine due to drug metabolism, as expected from each compound's pharmacology. A computational fluid dynamics simulation enabled the prediction of terfenadine-fexofenadine pharmacokinetics, which was validated by HPLC-MS. This in vitro platform recapitulates primary aspects of the in vivo crosstalk between heart and liver and enables pharmacological studies, involving both organs in a single in vitro platform. The system enables non-invasive readouts of cardiotoxicity of drugs and their metabolites. Hepatotoxicity can also be evaluated by biomarker analysis and change in metabolic function. Integration of metabolic function in toxicology models can improve adverse effects prediction in preclinical studies and this system could also be used for chronic studies as well.

The immune system as a chronotoxicity target of the anticancer mTOR inhibitor everolimus.

The circadian timing system controls many biological functions in mammals including xenobiotic metabolism, detoxification, cell proliferation, apoptosis and immune functions. Everolimus is a mammalian target of rapamycin inhibitor, whose immunosuppressant properties are both desired in transplant patients and unwanted in cancer patients, where it is indicated for its antiproliferative efficacy. Here we sought whether everolimus circadian timing would predictably modify its immunosuppressive effects so as to optimize this drug through timing. C57BL/6J mice were synchronized with light-dark 12h:12h, with L onset at Zeitgeber Time (ZT) 0. Everolimus was administered orally to male (5 mg/kg/day) and female mice (15 mg/kg/day) at ZT1, during early rest span or at ZT13, during early activity span for 4 weeks. Body weight loss, as well as hematological, immunological and biochemical toxicities, were determined. Spleen and thymus were examined histologically. Everolimus toxicity was less severe following dosing at ZT13, as compared to ZT1, as shown with least body weight inhibition in both genders; least reductions in thymus weight both in males (p < 0.01) and females (p < 0.001), least reduction in female spleen weight (p < 0.05), and less severe thymic medullar atrophy both in males (p < 0.001) and females (p < 0.001). The mean circulating counts in total leukocytes, total lymphocytes, T-helper and B lymphocytes displayed minor and non-significant changes following dosing at ZT13, while they were decreased by 56.9% (p < 0.01), 45.5% (p < 0.01), 43.1% (p < 0.05) and 48.7% (p < 0.01) after everolimus at ZT1, respectively, in only male mice. Chronotherapy of everolimus is an effective way to increase the general tolerability and decrease toxicity on the immune system.

Yupingfeng Powder relieves the immune suppression induced by dexamethasone in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Yupingfeng Powder (YPF), a Chinese medical formula, is used traditionally for allergic diseases and characterized by reducing allergy relapse. In the present study, we attempted to investigate the effect of YPF on the immunity of mice and the possible mechanisms. MATERIALS AND METHODS: An immunosuppressive mice model induced by Dexamethasone (Dex) was used. Blood samples, spleen and thymus were collected. Then, hematology parameters and organ weight were measured; Phenotypic analyses (CD4+/CD8+) of lymphocytes were performed using a flow cytometer; Phagocytosis of peritoneal macrophages were evaluated by particle tracers; Spleen lymphocytes were isolated, whose proliferation and apoptosis were assessed. NK cells' cytotoxicity was determined using the LDH release assay. RESULTS: YPF could ameliorate weight loss and improve low thymus and spleen coefficients caused by Dex. Treatment with YPF made decreased lymphocytic activity of Dex-treated mice back to normal and inhibited Dex-induced apoptosis of lymphocytes. YPF increased the Dex caused low proportion of CD4+/CD8+, and upregulated Dex-reduced NK cells' activity. CONCLUSION: The series of experiments demonstrated that YPF could exert immune regulation and enhance immunity of immunosuppressive mice through adjusting nonspecific and cellular immunity. The results would provide a basis for clinical application of YPF.

A combined approach to early detect in vitro drug-induced hemostatic changes in preclinical safety.

Early detection of drug-induced alterations of hemostasis is challenging. Drugs can affect different components of the Virchow's triad and measurement of plasmatic coagulation times lacks sensitivity. New techniques for a more global assessment of the hemostasis are now available: the impedance platelet aggregometry, the thromboelastography and the thrombin generation measurement. The aim of this study was to evaluate three techniques (i.e.: Multiplate®, TEG® and CAT) for the in vitro detection of the effect of a drug known to induce hemostatic alterations in a preclinical safety environment. Cyclosporine A was chosen and tested at 4 concentrations after solubilization in DMSO in Wistar rats and Beagle dogs. The results obtained were comparable between both species except for the thrombin generation in platelet rich plasma. Enhanced platelet aggregability was observed after ADP stimulation and alterations of the thromboelastograms consisted in decreased maximum amplitude and increased LY30. A dual effect on thrombin generation was observed and suggested that CsA may interact with platelets in rat platelet rich plasma and speed up thrombin generation. The results of this study indicate that using a combined approach on hemostasis testing in preclinical safety it is possible to detect in vitro drug-induced alterations of hemostasis.