Renal Protective Effects of Inonotus obliquus on High-Fat Diet/Streptozotocin-Induced Diabetic Kidney Disease Rats: Biochemical, Color Doppler Ultrasound and Histopathological Evidence

Diabetic kidney disease (DKD) is the current leading cause of end-stage renal disease. Inonotus obliquus (chaga), a medicinal fungus, has been used in treatment of diabetes. Here, we aim to identify the renal protective effects of chaga extracts on a DKD rat model which was induced by a high-fat diet and streptozotocin injection. During the total 17-weeks experiment, the biological parameters of serum and urine were examined, and the color Doppler ultrasound of renal artery, the periodic acid-Schiff staining, and electron microscopy of kidney tissue were performed. The compositions of chaga extracts were analyzed and the intervention effects of the extracts were also observed. Compared with the normal control group, the biochemical research showed that insulin resistance was developed, blood glucose and total cholesterol were elevated, urinary protein excretion and serum creatinine levels were significantly increased in the DKD model. The ultrasound examinations confirmed the deteriorated blood flow parameters of the left renal interlobar artery in the rat models. Finally, histopathological data supported renal injury on the thickened glomerular basement membrane and fusion of the foot processes. 8 weeks intervention of chaga improved the above changes significantly, and the 100 mg/kg/d chaga group experienced significant effects compared with the 50 mg/kg/d in some parameters. Our findings suggested that Doppler ultrasound examinations guided with biochemical indicators played important roles in evaluating the renal injury as an effective, noninvasive, and repeatable method in rats. Based on biochemical, ultrasound, and histopathological evidence, we confirmed that chaga had pharmacodynamic effects on diabetes-induced kidney injury and the aforementioned effects may be related to delaying the progression of DKD.

Inonotus obliquus. [Descriptions of Fungi and Bacteria].

A description is provided for Inonotus obliquus, which forms black circular or irregular brittle charcoal- or clinker-like crusts or conks, breaking out of bark on trunks of living trees, particularly Betula species. Some information on its dispersal and transmission and conservation status is given, along with details of its geographical distribution (Greenland, Armenia, China (Shanxi), India (Uttarakhand) Iran, Japan, Kazakhstan (Almaty, East Kazakhstan, Kostanay, West Kazakhstan), Mongolia, Nepal, Russia (Altai Krai, Amur Oblast, Kamchatka Krai, Khanty-Mansi Autonomous Okrug, Krasnoyarsk Krai, Novosibirsk Oblast, Omsk Oblast, Primorsky Krai, Republic of Altai, Sakhalin Oblast, Sverdlovsk Oblast, Tomsk Oblast, Tyumen Oblast, Yamalo-Nenets Autonomous Okrug), Sri Lanka, Uzbekistan, Austria, Belarus, Bulgaria, Czech Republic, Denmark, Estonia, Finland, France, Germany, Hungary, Ireland, Italy, Latvia, Lithuania, Netherlands, Norway, Poland, Romania, Russia (Arkhangelsk Oblast, Bryansk Oblast, Chuvash Republic, Ivanovo Oblast, Kaliningrad Oblast, Kaluga Oblast, Kirov Oblast, Komi Republic, Kostroma Oblast, Kursk Oblast, Leningrad Oblast, Moscow Oblast, Murmansk Oblast, Nizhny Novgorod Oblast, Novgorod Oblast, Orenburg Oblast, Pskov Oblast, Republic of Bashkortostan, Republic of Karelia, Republic of Mordovia, Republic of Tatarstan, Smolensk Oblast, Tula Oblast, Tver Oblast, Udmurt Republic, Ulyanovsk Oblast, Vladimir Oblast, Vologda Oblast, Voronezh Oblast, Yaroslavl Oblast), Slovenia, Spain, Sweden, Ukraine, UK, Canada (Alberta, British Columbia, Manitoba, New Brunswick, Newfoundland and Labrador, Northwest Territories, Nova Scotia, Ontario, Prince Edward Island, Quebec, Saskatchewan, Yukon), USA (Alabama, Alaska, Connecticut, Florida, Idaho, Indiana, Kansas, Kentucky, Louisiana, Maine, Massachusetts, Michigan, Minnesota, Mississippi, Montana, New Hampshire, New Jersey, New York, North Carolina, Ohio, Pennsylvania, Rhode Island, South Carolina, Tennessee, Vermont, Virginia, Washington, West Virginia, Wisconsin)) and hosts (Betula spp. and Alnus spp.).

Inhibitory Effects of Inonotus obliquus Polysaccharide on Inflammatory Response in Toxoplasma gondii-Infected RAW264.7 Macrophages

Our previous reports have shown that Inonotus obliquus polysaccharide (IOP) has protective effects against Toxoplasma gondii (T. gondii) infection in vivo. The aim of the present research is to explore the in vitro anti-inflammatory effects of IOP and its mechanism in RAW264.7 macrophages infected by T. gondii. In this study, it is indicated that IOP decreased the excessive secretion of inflammatory cytokines tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-1β (IL-1β), IL-4, and IL-6 in T. gondii-infected RAW264.7 macrophages. IOP effectively suppressed the mRNA expression of these cytokines and chemokines monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1α (MIP-1α). Moreover, IOP inhibited the phosphorylation of inhibitor kappa B kinase α/β (IKKα/β), inhibitor κBα (IκBα), p65 in nuclear factor-kappa B (NF-κB) signaling pathway and p38, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase 1/2 (ERK1/2) in mitogen-activated protein kinases (MAPKs) signaling pathway. Meantime, IOP prevented NF-κB p65 and c-Jun translocation from the cytoplasm to the nucleus. Further, IOP downregulated the protein expression of toll-like receptor 2 (TLR2) and TLR4 in T. gondii-infected RAW264.7 macrophages. The above results suggest that IOP can inhibit the inflammatory response infected with T. gondii via regulating TLR2/TLR4-NF-κB/MAPKs pathways and exerting its anti-T. gondii role in vitro.

Lipid-Lowering Effects of Inonotus obliquus Polysaccharide In Vivo and In Vitro

Excessive lipid intake will cause hyperlipidemia, fatty liver metabolism disease, and endanger people’s health. Edible fungus polysaccharide is a natural active substance for lipid lowering. In this study, the HepG2 cell model induced by oleic acid and mice model induced by a high-fat diet was established. The lipid-lowering effects of Inonotus obliquus polysaccharide (IOP) was investigated in vivo and in vitro. Glucose (251.33 mg/g), rhamnose (11.53 mg/g), ribose (5.10 mg/g), glucuronic acid (6.30 mg/g), and galacturonic acid (2.95 mg/g) are present in IOP, at a ratio of 85.2:3.91:1.73:2.14:1. The molecular weight of IOP is 42.28 kDa. Treatment with 60 mg/L of IOP showed a significant lipid-lowering effect in HepG2 cells compared with the oleic acid-treated group. In the oil red O-stained images, the red fat droplets in the IOP-treated groups were significantly reduced. TC and TG levels of IOP-treated groups decreased. IOP can alleviate the lipid deposition in the mice liver due to high-fat diet, and significantly reduce their serum TC, TG, and LDL-C contents. IOP could activate AMPK but decrease the SREBP-1C, FAS, and ACC protein expression related to adipose synthesis in mice. IOP has a certain potential for lipid-lowering effects both in vivo and in vitro.

Inonotus obliquus polysaccharide ameliorates serum profiling in STZ-induced diabetic mice model

Background Diabetes mellitus is a systemic disease mainly caused by the disorder of metabolism, which has become huge threat to human health. Polysaccharides are the main active substance from Inonotus obliquus (I. obliquus) with hypoglycemic effect. This study aims to evaluate the hypoglycemic activity and investigate the molecular mechanism of I. obliquus polysaccharide (IOP) in streptozotocin (STZ)-induced diabetic mice using metabolomics based on UPLC-Q-Exactive-MS method. Results The results showed that the oral administration of IOP in high dose (1.2 g/kg) can significantly reduce the blood glucose with 31% reduction comparing with the diabetic model and relieve dyslipidemia in diabetic mice. By UPLC-Q-Exactive-MS method and multivariate statistical analysis, a total of 15 differential metabolites were identified, including 4 up-regulated and 11 down-regulated biomarkers, of which l-tryptophan, l-leucine, uric acid, 12-HETE, arachidonic acid, PC(20:1(11Z)/14:1(9Z)) and SM(d18:0/24:1(15Z)) were exhibited an important variation, as the potential biomarkers in diabetes. Pathway analysis indicated that phenylalanine, tyrosine and tryptophan biosynthesis and arachidonic acid metabolism were prone to interference in diabetes. Moreover, leucine and proline were reversed and phytosphingosine was further reduced in diabetic mice under the intervention of IOP. Conclusion IOP has predominant hyperglycemic effect on STZ-induced diabetic mice via ameliorating serum profiling.

Inonotus obliquus aqueous extract suppresses carbon tetrachloride induced hepatic injury through modulation of antioxidant enzyme system and anti-inflammatory mechanism

Background: Chaga mushroom [Inonotus obliquus] is an edible macrofungus used in traditional and folk medicine for treatment of various gastrointestinal disorders. It has shown potent anti-inflammatory, antioxidant and anticancer effects in several experimental studies including our anti-inflammatory and anticancer effects in colorectal cancer and intestinal inflammation. Whole extract or purified compound ergosterol peroxide from chaga mushroom showed anti-inflammatory mechanism via suppression of NF-κB/iNOS-COX-2 and growth inhibitory mechanism via regulation of apoptosis activation and β-catenin suppression. The emergence of diverse inflammatory and carcinogenic agents like carbon tetrachloride [CCl4] is a potent hepatotoxic chemical that caused liver damage by inducing lipid peroxidation and other oxidative damages. Aims: The study was aimed to analyze the biochemical, cellular and molecular mechanism of CCl4 induced chronic liver inflammation and carcinoma and to analyze the effect of the extract of chaga mushroom on liver inflammation and cancer by virtue of anti-inflammatory mechanisms. Method: Physiological, histological and immunohistochemical the physiological functions and cellular functions. Biochemical assays for assessing enzymatic changes in tissues. Molecular simulation and docking studies were performed for proposing the molecular interaction. Results: CCl4-exposed mice exhibited a significant decrease in the body weight followed by altered histopathological signatures in the liver. Supplementation of IOAE showed that treatment restored towards normal structure of the tissues with large round nuclei in most of the cells. CCl4 caused a steep elevation in the levels of SGOT and SGPT to 2.32- and 1.8-fold as compared to control. The LDH level was increased to 447 IU/L in CCl4 treated mice as compared to control [236 IU/L]. Analysis of the oxidant enzyme pathway showed that CCl4 reduced the GSH level to 16.5 μM as compared to control [52 μM], and induced the catalase enzyme activity to 259 U/mL as compared to control [124 U/L]. These physiological and biochemical alterations were restored towards normal levels by IOAE administration. Immunohistochemical staining for caspase-3 and p53 showed that CCl4 notably increased their expressions which were subsequently suppressed by administration of IOAE. The molecular simulation and docking studies using ergosterol peroxide from chaga mushroom with iNOS, COX-2 and TNF-α showed binding energy of -10.5, -8.9 and -9.1 Kcal/mol, respectively. These proteins interacting with ergosterol peroxide suggests an inhibitory effect on these critical proinflammatory signaling proteins. Conclusions: The results point out that IOAE is able to prevent damage of hepatic cells caused by CCl4 in mouse models through anti-inflammatory and growth inhibitory mechanism which can be utilized in natural prevention of the liver toxicity.

Research of Inonotus obliquus Oligosaccharide in Prevention of Hyperlipidemia

In this study, hot water was used to extract Inonotus obliquus oligosaccharide. DEAE-cellulose and Sepharose G-200 were used to purify Inonotus obliquus oligosaccharide. Inonotus obliquus oligosaccharide IOP-2A was obtained. Its molecular weight Mw is about 1000 Da. The monosaccharide composition and molar ratio were glucose : xylose : galactose : mannose = 54.1 : 13.6 : 13.2 : 6.7. In addition, it also contains a small amount of galactose, gluconic acid, rhamnose, and fucose. IOP-2A contained mainly β-glycosidic bonds. Among them, 1,4-glycosidic bonds accounted for 9.2%, and 1,6-glycosidic bonds accounted for 85.1%. Oligosaccharide macromolecules formed a layered structure. Mouse experiments showed that IOP-2A had the function of preventing hyperlipidemia. At the same time, IOP-2A had a certain protective effect on the liver and kidney. The mechanism of IOP-2A in preventing hyperlipidemia was obtained from the perspective of mouse intestinal flora.

Convergent evolution of mevalonate pathway in Inonotus obliquus and Betula pendula.

Inonotus obliquus, Chaga mushroom, is a fungal species from Hymenochaetaceae family (Basidiomycota) which has been widely used for traditional medicine in Europe and Asia. Here, chaga genome was sequenced using Pacbio sequencing into a 50.7Mbp assembly consisting of 301 primary contigs with an N50 value of 375 kbp. Genome evolution analyses revealed a lineage-specific whole genome duplication event and an expansion of Cytochrome P450 superfamily. Fungal biosynthetic clusters were enriched for tandemly duplicated genes, suggesting that biosynthetic pathway evolution has proceeded through small-scale duplications. Metabolomic fingerprinting confirmed a highly complex terpene biosynthesis chemistry when compared against related fungal species lacking the genome duplication event.