Apoptosis In Liver Research Articles

Hesperidin ameliorates liver ischemia/reperfusion injury via activation of the Akt pathway.

Hesperidin (HDN) is a bioflavonoid that serves a role as an antioxidant in biological systems. However, although HDN has hydrogen radical- and hydrogen peroxide-removal activities, the role of HDN in liver ischemia/reperfusion (I/R) injury remains unknown. This study aimed to determine the role of HDN in liver I/R injury. Male C57BL/6J wild-type (WT) mice were subjected to warm partial liver I/R injury. Liver damage was evaluated by measuring serum alanine aminotransferase (ALT) levels, cytokine production, oxidative stress indicators, tissue hematoxylin-eosin staining and cell death. The Akt signaling pathway was examined to elucidate the underlying mechanisms. HDN had no effect on ALT levels and tissue damage in WT mice without liver I/R injury. However, HDN significantly ameliorated liver I/R injury as measured by serum ALT levels and necrotic tissue areas. HDN decreased malondialdehyde content, but increased the levels of superoxide dismutase, catalase, glutathione peroxidase and glutathione. In addition, HDN significantly attenuated the mRNA expression levels of TNF-α, IL-6 and IL-1β after liver I/R injury. Furthermore, HDN protected the liver against apoptosis in liver I/R injury by increasing the levels of Bcl-2 and decreasing the levels of cleaved-caspase 3. Mechanistically, the levels of phosphorylated Akt were elevated by HDN during liver I/R injury. In addition, HDN could induce Akt activation in hepatocytes in vitro. Most importantly, treatment with the Akt inhibitor LY294002 in WT mice blocked the hepatoprotective effects of HDN in liver I/R injury. In summary, the results of the present study suggested that HDN may protect against liver I/R injury through activating the Akt pathway by ameliorating liver oxidative stress, suppressing inflammation and preventing hepatocyte apoptosis. HDN may be a useful factor for liver injury protection and a potential therapeutic treatment for liver I/R injury in the future.

Induction of Surgical Obstructive Cholestasis in rats: morphological, biochemical and immunohistochemical changes

Objective: To detail an easy approach for surgical induction of obstructive cholestasis in a rat model. Additionally, to assess the role of serum and tissue biomarkers for determination of the extent of cholestasis induced hepatocellular injury and to quantify the apoptotic liver cell using fluorescein-labeled Annexin V. Design: Randomized controlled experimental study. Animals: Sixty-five healthy Male Sprague- Dawley rats weighing 268.18 ± 23.3 grams. Procedures: The rats were randomly allocated into two main groups: 1) sham group (SO; n= 15) and 2) bile duct ligation group (BL; n= 50). Three SO rats and 8 BL rats were euthanized at 10, 20, 30, 45 and 60 post-surgery. Blood and liver samples were collected for serum and tissue biochemical analysis. Liver apoptosis was quantified using Annexin V, meanwhile liver fibrosis was quantified using both TNF-β and histological examination. Results: There was a tendency towards less complications and higher surgeon satisfaction when the common bile duct was approached by the stomach approach. Serum biochemical analysis revealed a significant time-dependent decreases in hepatic serum biomarkers (alanine aminotransferease, aspartate aminotransferase, serum glutamic oxaloacetic transaminase, alkaline phosphatase, gamma-glutamyltransferase, serum albumin, the concentration of total bilirubin, and direct bilirubin) after induced obstructive cholestasis (P = 0.01-0.001), whereas AST: ALT ratio conversely was elevated (p < 0.05 value). Flow cytometer analysis revealed time-dependent significant increase in the percent of apoptotic cells after BL (P < 0.001) and consequently the state of oxidative stress (malondialdehyde) was (P < 0.001). The antioxidant tissue markers (superoxide dismutase, Catalase and glutathione reductase) were significantly decreased (P < 0.05), meanwhile TNF-β, the fibrosis marker showed time dependent significant increases (P = 0.001). Conclusion and clinical relevance: Stomach approach provided an easy, fast and reliable procedure for induction of obstructive cholestasis in rats. Liver Annexin V and TGF-β were diagnostic tools in further apoptosis and fibrosis.

The protective effect of Morin against ifosfamide-induced acute liver injury in rats associated with the inhibition of DNA damage and apoptosis

Morin is a flavonoid and broadly found in white berry and cranberry branch. Ifosfamide (IFOS) is known as an anticancer and cytotoxic drug especially on the liver. This study aimed to explore the potential protective effects of Morin against IFOS-induced liver toxicity in rats. The model group of rats received a single injection of IFOS (500 mg/kg; i.p.) at day 2, whereas the protective groups of rats were given two different doses of Morin (100 and 200 mg/kg; given by gavage) at days 1 and 2. All animals were then culled 24 h post-IFOS injection. We observed that IFOS caused liver injury, oxidative stress, inflammation, DNA damage, and apoptosis. However, Morin decreased the levels of aspartate aminotransferase (AST), alkaline phosphatase (ALP), alanine aminotransferase (ALT) (p < 0.05). While Morin contributed to the recovery of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione (GSH) levels, Morin decreased the levels of malondialdehyde (MDA) induced by IFOS in the liver (p < 0.05). Besides, the levels of nuclear factor kappa B (NF-κB), tumor necrosis factor-α (TNF-α), and P53 measured by ELISA test were reduced via Morin administration (p < 0.05). Lastly, the mRNA transcript levels of Bax, Apaf-1, Bcl-2, Bcl-xL, and inducible nitric oxide synthase (iNOS) determined by RT-PCR were down-regulated in the Morin groups (p < 0.05). These results indicate that Morin plays a protective role by reducing oxidative stress, inflammation, and apoptosis in the IFOS-induced liver injury in rats.

Hematological and Hepatic Effects of Ulexite in Zebrafish

The ulexite (UX), a borate mineral, is used as boron source and commonly used in various industrial processes. The hematological and hepatic effects of UX were investigated by exposing adult zebrafish to UX (5, 10, 20 and 40 mg/L) over 96 hours. The blood and liver tissues were taken at the end of the trial period then micronucleus (MN) rates, oxidative DNA damage (8-OHdG), apoptosis (Caspase-3), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), myeloperoxidase (MPO), paraoxonase (PON), arylesterase (AR) and lipid peroxidation (MDA) levels were determined. Genotoxic damage by UX occurred only at 40 mg/L in the blood MN assay. Oxidative stress, oxidative DNA damage and apoptosis in liver also occurred at this dose. Moreover, 5-20 mg/L doses led to decreases of DNA damage and apoptosis levels via promoting antioxidant system in liver tissues. UX exhibits beneficial roles on blood and liver tissues of zebrafish at relatively lower doses, which may be relevant to nutritional and medicinal industries.

Kupffer cell inactivation ameliorates immune liver injury via TNF-α/TNFR1 signal pathway in trichloroethylene sensitized mice

Aim: The present study aims to explore the important role of Kupffer cell and its related cytokine TNF-α in immune liver injury induced by trichloroethylene (TCE). Methods 36 female BALB/c mice were selected and randomly divided the mice into four groups. We established a BALB/c mouse model of TCE sensitization and pretreatment with GdCl3 (40 mg/kg) by intraperitoneal injection during the during the 17th and 19th days. Results We found F4/80, the marker of Kupffer cell, was increased in TCE positive group. GdCl3 treatment successfully blocked the activation of Kupffer cell. TNF-α was increased significantly in liver of TCE sensitized mice and decreased significantly when low-dose GdCl3 was used. We found TNF receptor 1 (TNFR1) was increased significantly and GdCl3 treatment resumed the expression of TNFR1 to normal level, as well as the F4/80, TNF-α and TNFR1 mRNA. We also found both caspase-8 and caspase-3 increased in TCE positive group and decreased in TCE + GdCl3 positive group. The number of apoptotic cells in TCE sensitized mice increased by TUNEL staining, and GdCl3 treatment alleviated this increase. Some cells showed edema and inflammatory cell aggregation in liver of TCE positive group, while in the TCE + GdCl3 positive group, the cytoplasm became loose and vacuole-like degeneration occurred. Conclusion Our study unveils cross-talk between Kupffer cell activation and TNFR1 which mediate apoptosis in liver of TCE sensitized mice.

Effect of acupuncture on oxidative stress and apoptosis-related proteins in obese mice induced by high-fat diet

To explore the effect of acupuncture on oxidative stress and apoptosis-related proteins of liver in obese mice induced by high-fat diet. A total of 45 male C57BL/6 mice were randomized into a control group (10 mice) and a model established group (35 mice). Mice in the model established group were fed with high-fat diet for 16 weeks to establish the obesity model. After model established, 30 mice were randomized into a model group, a non-acupoint group and an acupoint group, 10 mice in each one. Acupuncture was applied at "Guanyuan" (CV 4), " Zusanli" (ST 36), "Yishu" (EX-B 3) in the acupoint group and the points of 0.5 cm and 1 cm to the base of tail in the non-acupoint group, 15 min each time, once a day for 8 weeks. Mice in the control group were fed with normal diet, while mice in the other 3 groups were fed with high-fat diet continuously for 8 weeks. The body weight was measured at 0, 4th, 8th, 12th, 16th, 20th, 24th week in each group respectively. After 24-week intervention, the weight of white adipose tissue of epididymis and perirenal and liver was measured; the levels of serum alanine transaminase(ALT) and glutamic oxaloacetic aminotransferase (AST) were detected by automatic biochemical analyzer; liver homogenate was used to detect the level of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD); the liver morphology was observed by HE staining; the expression of apoptosis-related proteins Bax and Bcl-2 were detected by Western blot. Compared with the control group, the body weight of mice in the model group, the acupoint group and the non-acupoint group was decreased on 16th week into experiment (before intervention, P<0.05); compared with the model group and the non-acupoint group, the body weight of mice in the acupoint group were decreased after intervention (P<0.05). Compared with the control group, the weight of white adipose tissue and liver, the levels of serum ALT and AST, the level of liver MDA, the expression of liver Bax were increased (P<0.05); the activity of liver SOD and the expression of liver Bcl-2 were decreased in the model group after intervention (P<0.05). Compared with the model group and the non-acupoint group, the weight of white adipose tissue and liver, the levels of serum ALT and AST, the level of liver MDA, the expression of liver Bax were decreased (P<0.05); the activity of liver SOD and the expression of liver Bcl-2 were increased in the acupoint group after intervention (P<0.05). Acupuncture at "Guanyuan" (CV 4), "Zusanli" (ST 36) and "Yishu"(EX-B 3) can improve obesity and obesity related hepatic disorder by regulating oxidative stress and inhibiting apoptosis in liver.

Tannic acid attenuates hepatic oxidative stress, apoptosis and inflammation by activating the Keap1‑Nrf2/ARE signaling pathway in arsenic trioxide‑toxicated rats

The present study was performed to investigate the protective effects of tannic acid (TA) on liver injury induced by arsenic trioxide (ATO) and to elucidate the mechanism involved as related to the Kelch‑like ECH‑associated protein 1 (Keap1)‑nuclear factor erythroid 2‑related factor2 (Nrf2)/antioxidant response element (ARE) signaling pathway. Adult rats were intraperitoneally injected with TA, while ATO was administered 1h later. On the 11th day, the rats were euthanized to determine any liver histological changes, liver function, and the activities of antioxidant, antiapoptosis and proinflammatory cytokines in the liver. Furthermore, the protein expression levels of nuclear Nrf2, total Nrf2, Keap1, Heme oxygenase‑1 (HO‑1), NADPH quinine oxidoreductase‑1 (NQO1), and γ‑glutamylcysteine synthetase (γ‑GCS) were determined using western blot analysis. The results showed that TA treatment ameliorated ATO‑induced liver histological changes and decreased the ATO‑induced increased alanine aminotransferase (ALT) and aspartate transaminase (AST) serum levels. Activities of the antioxidant enzymes significantly were increased, while the levels of malondialdehyde (MDA) and reactive oxygen species (ROS) were attenuated following TA treatment. In addition, TA treatment inhibited ATO‑induced liver apoptosis and inflammatory responses, increased Bcl‑2 protein expression level and reduced the levels of Bax, caspase‑3, interleukin (IL)‑1β, IL‑6 and tumor necrosis factor (TNF)‑α. Furthermore, TA treatment increased the protein expression levels of Nrf2 and Keap1, HO‑1, NQO1 and γ‑GCS. The results demonstrated that TA has a protective effect on ATO‑treated hepatic toxicity and that its underlying mechanism could be due to TA activation of the Keap1‑Nrf2/ARE signaling pathway, to reduce oxidative stress, apoptosis and inflammation in ATO‑intoxicated rats.

Molecular mechanism of olaquindox-induced hepatotoxicity and the hepatic protective role of curcumin

Olaquindox (OLA) is a chemosynthetic growth promoter, which could promote the treatment of bacterial infections and improve feed energy efficiency. Hepatotoxicity is still a poor feature associated with the adverse effects of OLA. The present study aimed to investigate the molecular mechanism of OLA-induced hepatotoxicity and the protective role of curcumin in mice and HepG2 cells. The result showed that representative biomarkers involved in mitochondrial pathway, p53 pathway, mitogen-activated protein kinase (MAPK) pathway, autophagy and antioxidant pathway were activated. Furthermore, curcumin attenuated OLA-induced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) and liver damage in mice. In addition, cell viability of HepG2 was enhanced by curcumin pretreatment at 5, 10 and 20 μM. Meanwhile, curcumin markedly ameliorated OLA-induced oxidative stress, apoptosis and mitochondrial dysfunction. Moreover, curcumin pretreatment significantly up-regulated the expressions of nuclear factor erythroid-2-related factor 2 (Nrf2) and heme oxygenase-1(HO-1) and down-regulated the expressions of nuclear factor-kappaB (NF-kB) and p53 through reduced the nuclear translocation of NF-kB induced by OLA. In summary, our findings indicated that OLA-induced hepatotoxicity involved in mitochondrial apoptosis, autophagy, p53 pathway, Nrf2/HO-1 pathways, and curcumin regulated OLA-induced liver damage, oxidative stress and apoptosis via activation of Nrf2/HO-1 pathway and suppression of p53 and NF-kB pathway.

Silibinin-hydroxypropyl-β-cyclodextrin (SLB-HP-β-CD) complex prevents apoptosis in liver and kidney after hepatic ischemia-reperfusion injury

BackgroundWe investigated the protective effect of silibinin on rat liver and kidney after hepatic inschemia/reperfusion (I/R) injury. Methods and materialsSixty three male Wistar-type rats (median age 13 weeks; average weight 314 g) were subjected to I/R injury of the liver. They were randomly divided into three groups: Sham (n = 7), Control (C, n = 28) and Silibinin (Si, n = 28). The last group received intravenously silibinin. The C and Si groups were each subdivided in four subgroups according to euthanasia times (i.e., 60, 120, 180, 240 min). We assessed expression of caspase-3 and TUNEL assay, and biochemical and histological parameters. ResultsAt 240 min, expression of caspase-3 and TUNEL assay were statistically significantly lower in the Si compared to the C group for both liver and kidney. SGOT and SGPT were also statistically significantly lower in the Si than in the C group at all time points. Histological parameters of the liver were also improved in the Si group. ConclusionSilibinin was found to exhibit a protective effect on liver and kidney after hepatic I/R injury. The present results are encouraging for further studies and future clinical application.

Artemisinin Derivatives Stimulate DR5-Specific TRAIL-Induced Apoptosis by Regulating Wildtype P53.

Simple SummaryThe risk of developing colorectal cancer at a younger age has increased, but current therapies are either risky or limited. We aim to demonstrate that the combination treatment of artemisinin derivatives and TRAIL could be a potential therapy to kill colon cancer cells. We found that artemisinin derivatives increase death receptor production and further sensitize colon cancer cells to TRAIL-induced apoptosis. Furthermore, we explored the role of P53 in response to artemisinin derivatives, which transactivates Death Receptor 5 (DR5) and the cyclin-dependent kinase inhibitor P21. Finally, a 3D tumor spheroid model also confirmed the efficacy of the combination treatment.Artemisinin derivatives, widely known as commercial anti-malaria drugs, may also have huge potential in treating cancer cells. It has been reported that artemisinin derivatives can overcome resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in liver and cervical cancer cells. In our study, we demonstrated that artesunate (ATS) and dihydroartemisinin (DHA) are more efficient in killing colon cancer cells compared to artemisinin (ART). ATS/DHA induces the expression of DR5 in a P53 dependent manner in HCT116 and DLD-1 cells. Both ATS and DHA overcome the resistance to DHER-induced apoptosis in HCT116, mainly through upregulating death receptor 5 (DR5). We also demonstrate that DHA sensitizes HCT116 cells to DHER-induced apoptosis via P53 regulated DR5 expression in P53 knockdown assays. Nevertheless, a lower effect was observed in DLD-1 cells, which has a single Ser241Phe mutation in the P53 DNA binding domain. Thus, the status of P53 could be one of the determinants of TRAIL resistance in some cancer cells. Finally, the combination treatment of DHA and the TRAIL variant DHER increases cell death in 3D colon cancer spheroid models, which shows its potential as a novel therapy.

Induction of Surgical Obstructive Cholestasis in rats: morphological, biochemical and immunohistochemical changes

Objective: To detail an easy approach for surgical induction of obstructive cholestasis in a rat model. Additionally, to assess the role of serum and tissue biomarkers for determination of the extent of cholestasis induced hepatocellular injury and to quantify the apoptotic liver cell using fluorescein-labeled Annexin V. Design: Randomized controlled experimental study. Animals: Sixty-five healthy Male Sprague- Dawley rats weighing 268.18 ± 23.3 grams. Procedures: The rats were randomly allocated into two main groups: 1) sham group (SO; n= 15) and 2) bile duct ligation group (BL; n= 50). Three SO rats and 8 BL rats were euthanized at 10, 20, 30, 45 and 60 post-surgery. Blood and liver samples were collected for serum and tissue biochemical analysis. Liver apoptosis was quantified using Annexin V, meanwhile liver fibrosis was quantified using both TNF-β and histological examination. Results: There was a tendency towards less complications and higher surgeon satisfaction when the common bile duct was approached by the stomach approach. Serum biochemical analysis revealed a significant time-dependent decreases in hepatic serum biomarkers (alanine aminotransferease, aspartate aminotransferase, serum glutamic oxaloacetic transaminase, alkaline phosphatase, gamma-glutamyltransferase, serum albumin, the concentration of total bilirubin, and direct bilirubin) after induced obstructive cholestasis (P = 0.01-0.001), whereas AST: ALT ratio conversely was elevated (p < 0.05 value). Flow cytometer analysis revealed time-dependent significant increase in the percent of apoptotic cells after BL (P < 0.001) and consequently the state of oxidative stress (malondialdehyde) was (P < 0.001). The antioxidant tissue markers (superoxide dismutase, Catalase and glutathione reductase) were significantly decreased (P < 0.05), meanwhile TNF-β, the fibrosis marker showed time dependent significant increases (P = 0.001). Conclusion and clinical relevance: Stomach approach provided an easy, fast and reliable procedure for induction of obstructive cholestasis in rats. Liver Annexin V and TGF-β were diagnostic tools in further apoptosis and fibrosis.

Circ_0000105 promotes liver cancer by regulating miR-498/PIK3R1.

A growing number of studies demonstrate that circular RNA (circRNA) has a prominent role in the regulation of numerous biological behaviors of tumor cells. The present study aimed to investigate the expression, function and molecular mechanisms of circ_0000105 in liver cancer. A quantitative real-time polymerase chain reaction (qRT-PCR) was utilized to obtain the expressions of circ_0000105 and miR-498 in liver cancer tissues and cells. The association between the expression level of circ_0000105 and the clinicopathological characteristics of liver cancer patients was analyzed. qRT-PCR and western blotting were utilized to investigate the expression of phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1) in cells. Cell counting kit-8, bromodeoxyuridine and flow cytometry assays were utilized to determine liver cancer cell proliferation and apoptosis, respectively. Bioinformatics, a luciferase reporter gene experiment and a RNA immunoprecipitation experiment were conducted to predict and confirm the targeting correlation of circ_0000105 and miR-498, as well as miR-498 and PIK3R1. Circ_0000105 was highly expressed in liver cancer tissues and cell lines. Moreover, its high expression was associated with an increase in the T stage of liver cancer patients and a low degree of tumor differentiation. Circ_0000105 overexpression promoted liver cancer cell proliferation and inhibited apoptosis, whereas knocking down circ_0000105 triggered the opposite effect. miR-498 was a downstream target of circ_0000105, inhibiting liver cancer cell proliferation and promoting apoptosis. Mechanistically, circ_0000105 indirectly up-regulated the expression of PIK3R1 by adsorbing miR-498. Circ_0000105 plays a cancer-promoting role in liver cancer by regulating the miR-498/PIK3R1 axis.

Rutin-added diet protects silver catfish liver against oxytetracycline-induced oxidative stress and apoptosis

It is unknown whether the flavonoid rutin can protect the silver catfish liver in response to exposure to a known stressor, such as the prophylactic usage of the antimicrobial agent oxytetracycline. Thus, the current study aimed to assess the effect of rutin incorporation into the silver catfish diet formulation on oxytetracycline-induced liver oxidative stress and apoptosis. Fish were split into four groups as follows: control, rutin (1.5 g kg diet−1), oxytetracycline (0.1 g kg diet−1) and rutin+oxytetracycline (1.5 g kg diet−1 and 0.1 g kg diet−1, respectively). After two weeks of feeding with the different diets (standard, rutin-, oxytetracycline and rutin+oxytetracycline-added diets), fish were euthanized to collect the liver. Although the rutin-added diet was unable to recover glutathione peroxidase activity, ascorbic acid and reduced glutathione (GSH) levels, which were depleted due to oxytetracycline consumption, it markedly diminished the oxidized glutathione (GSSG) content, thus decreasing the GSSG to GSH ratio, an important index of oxidative stress. It also increased glutathione reductase and markedly augmented glucose-6-phosphate dehydrogenase activities, which were declined after oxytetracycline ingestion. Furthermore, the rutin-added diet reestablished superoxide dismutase and catalase activities and reduced lipid peroxidation, nitric oxide and superoxide anion levels as well, all changes resulting from oxytetracycline consumption. Finally, it also prevented oxytetracycline-induced apoptosis through increasing heat shock protein 70 and markedly decreasing high mobility group box 1 and, consequently, reducing cleaved caspase-3 protein levels. Therefore, in conclusion, the incorporation of this flavonoid to the silver catfish diet protected the liver against oxytetracycline-induced liver oxidative stress and apoptosis.

Sorafenib Affects Liver Cancer Cell Proliferation and Apoptosis by Regulating DJ-1-PTEN/PI3K/AKT Signaling Pathway

Objective: To assess sorafenib's role in regulating DJ-1-PTEN/PI3 K/AKT pathway and affecting the biological effects of liver cancer cells. Methods: Human normal liver HL-7702, liver cancer MHC97-L, and HCCLM3 cells were cultured in vitro to measure DJ-1 and PTEN expression. MHC97-L and HCCLM3 cells were treated with 0, 1.0, 2.0 M of sorafenib followed by analysis of cell viability by CCK-8, and DJ-1 and PTEN expressions by Western blot. MHC97-L cells were separated into control group, 2.0 M Sorafenib treatment group, Sorafenib + pcDNA3.1-Blank group, and Sorafenib + pcDNA3.1-DJ-1 group. Cell proliferation was assessed by flow cytometry and EdU staining. Results: Compared with HL-7702 cells, DJ-1 expression was significantly increased, while PTEN level was significantly declined in MHC97-L and HCCLM3 cells. Sorafenib treatment significantly inhibited the proliferation activity of MHC97-L and HCCLM3 cells. Different concentrations of sorafenib significantly downregulated DJ-1 expression and enhanced PTEN expression in MHC97-L cells. pcDNA3.1-DJ-1 transfection on the basis of sorafenib treatment significantly elevated DJ-1 and p -AKT levels, reduced PTEN expression, decreased cell apoptosis, and enhanced cell proliferation. Conclusion: Sorafenib downregulates DJ-1 expression and affects PTEN/PI3K/AKT pathway activity to exert an anti-tumor effect that inhibits liver cancer cell proliferation and promotes cell apoptosis.

Secondary Metabolites of Saussurea costus Leaf Extract Induce Apoptosis in Breast, Liver, and Colon Cancer Cells by Caspase-3-Dependent Intrinsic Pathway.

Background Apoptosis, a major form of programmed cell death, plays a vital role in regulating tissue development and maintenance of homeostasis in eukaryotes. Apoptosis can occur via a death receptor-dependent extrinsic or a mitochondrial-dependent intrinsic pathway and can be induced by various chemotherapeutic agents. In this study, the anticancer activity of Saussurea costus and its mode of intervention in human cancer cells of breast, colon, and liver were investigated. Results In this study, the bioactives of S. costus leaves were extensively extracted in five solvents of different polarity. The cytotoxicity and anticancer effect of the extracted secondary metabolites were investigated against breast (MCF-7), liver (HepG2), and colon (HCT116) cancer cell lines using a Sulphorhodamine B (SRB) assay. Secondary metabolites extracted using hexane, methanol, ethyl acetate, and chloroform had the highest cytotoxicity and thus the greatest anticancer effect on all the cancer cell lines tested (IC50; ranging from 0.25 to 2.5 μg/ml), while butanol was comparatively less active (IC50; ranging from 23.2 to 25.5 μg/ml). Further investigation using DNA flow cytometry and fluorescent microscopy revealed that the extract arrested the cells in the G1 phase of cell cycle and induced apoptosis. Furthermore, the elevated expression level of proapoptotic proteins and decreased expression level of antiapoptotic proteins confirmed that the intrinsic (mitochondrial) pathway was involved in mediating the apoptosis of cancer cells upon treatment with S. costus extract. These results altogether suggest that S. costus could be a potential anticancer agent. Conclusion These results suggest that the S. costus extract is the potential source of the secondary metabolites that could be used as anticancer agent to treat diverse cancers of breast, colon, and liver.

Chronic exposure to environmentally relevant concentration of fluoride alters Ogg1 and Rad51 expressions in mice: Involvement of epigenetic regulation

Chronic exposure to fluoride (F) beyond the permissible limit (1.5 ppm) is known to cause detrimental health effects by induction of oxidative stress-mediated DNA damage overpowering the DNA repair machinery. In the present study, we assessed F induced oxidative stress through monitoring biochemical parameters and looked into the effect of chronic F exposure on two crucial DNA repair genes Ogg1 and Rad51 having important role against ROS induced DNA damages. To address this issue, we exposed Swiss albino mice to an environmentally relevant concentration of fluoride (15 ppm NaF) for 8 months. Results revealed histoarchitectural damages in liver, brain, kidney and spleen. Depletion of GSH, increase in lipid peroxidation and catalase activity in liver and brain confirmed the generation of oxidative stress. qRT-PCR result showed that expressions of Ogg1 and Rad51 were altered after F exposure in the affected organs. Promoter hypermethylation was associated with the downregulation of Rad51. F-induced DNA damage and the compromised DNA repair machinery triggered intrinsic pathway of apoptosis in liver and brain. The present study indicates the possible association of epigenetic regulation with F induced neurotoxicity.

Ligustilide alleviates the insulin resistance, lipid accumulation, and pathological injury with elevated phosphorylated AMPK level in rats with diabetes mellitus

Background Diabetes mellitus (DM) is one of the major risk factors of disability and death worldwide. Despite of the protective role of ligustilide (LIG) in many cell types, we aimed to investigate whether LIG could be a potential to treat DM. Methods Sprague Dawley rats were randomly assigned to five groups. Rats except control were raised on a high-fat diet (HFD). Streptozotocin was intraperitoneally injected into HFD-fed rats to construct DM model. Rats in the LIG intervention groups received intraperitoneal injection of LIG (10, 20, and 40 mg/kg) post-induction of DM. Blood glucose, plasma insulin (p-insulin), adiponectin, HbA1C%, obesity index, HOMA-IR, and biochemical parameters were estimated. Histopathological analysis and apoptosis in liver and kidney, along with proliferation and apoptosis of islet β-cells, were analyzed. Expression of CPT-1 and ACC, and phosphorylation of Nrf2 and AMPKα1, were finally assessed. Results DM-induced alterations were all relived by LIG intervention. In brief, obesity index, glucose level, P-insulin content, HbA1C, and HOMA-IR were lowered while adiponectin level was elevated. Meanwhile, levels of TC, TG, ALT, and AST were decreased in the LIG intervention groups, along with up-regulated CPT-1 level and down-regulated ACC level. Pathological changes in liver and kidney tissues were alleviated, and apoptotic cells were reduced by LIG treatment. For islet β-cells, LIG up-regulated Ki67 and c-Myc expression, and mitigated ratios of Bax/Bcl-2 and cleaved cas3(9)/cas3(9). Finally, LIG could promote phosphorylation of Nrf2 and AMPKα1. Conclusions LIG alleviated the insulin resistance, lipid accumulation, and pathological injury with the activation of AMPK pathway in DM rats.

The assembly of glycosphingolipid determines their immunomodulatory effect: A novel method for structure-based design of immunotherapy

Structure-activity relationships provide insight into the binding interactions of beta-glycosphingolipids (GSLs) with both the TCR and the CD1d molecules, as well as the subsequent immunologic response of regulatory NKT cells. AimTo determine the effects of synthetic GSL structures on their immune modulatory functions. MethodsGSLs of various structures were tested in vitro and in an animal model of Concanavalin A (ConA) immune-mediated hepatitis. ResultsIn vitro, using SV40 binding to live monkey CV1 cells, the l-threo stereoisomer of C8-β-LacCer inhibits caveolar internalization, reducing viral binding to the cell surface. In vivo, in the ConA model, LR172, which has a saturated C8 chain, and LR178, which has a trans double bond at C-2 in the C8 chain, suppressed the immune-mediated liver inflammation and reduced IFNγ levels in a dose dependent manner. The beneficial effects of LR172 and of LR178 are associated with suppression of liver apoptosis, increased phosphorylated STAT3 expression in the liver, and an increase in the NKT liver/spleen ratio. SummaryThe assembly of GSLs determines their immunomodulatory effect and can serve as a method for structure-based design of immunotherapy.

Salidroside alleviated hypoxia-induced liver injury by inhibiting endoplasmic reticulum stress-mediated apoptosis via IRE1α/JNK pathway

Endoplasmic reticulum (ER) stress and subsequent apoptosis played vital role in liver injury and dysfunction. The aim of this study was to investigate the protective effect and mechanism of salidroside on hypoxia induced liver injury both in vivo and in vitro. Male SD rats were exposed to hypobaric chamber to simulate high altitude hypoxia model. High altitude hypoxia led to significant liver injury and apoptosis, increased the expression levels of p-JNK, BAX and ER stress markers. Salidroside treatment significantly inhibited hypoxia induced ER stress by decreasing the protein expression of glucose-regulated protein 78 (GRP78), CCAAT/enhancer binding protein homologous protein (CHOP) and phosphorylated inositol-requiring enzyme 1α (p-IRE1α). In addition, salidroside treatment also restrained the ER stress-mediated apoptotic pathway, as indicated by decreased pro-apoptotic proteins p-JNK, TRAF2, BAX, and cleaved caspase 9 and caspase 12, as well as upregulation of Bcl-2. Furthermore, in vitro study found that blocking IRE1α pathway using specific inhibitor STF-083010 subsequently reversed the protective effect of salidroside on liver apoptosis. Taken together, our findings revealed that salidroside exerts protective effects against hypoxia induced liver injury through inhibiting ER stress mediated apoptosis via IRE1α/JNK pathway.

Pharmaceutical efficacy of harmalol in inhibiting hepatocellular carcinoma

BackgroundDiethylnitrosamine (DEN) promoted by carbon tetrachloride (CCl4) forms DNA adducts inducing hepatocellular carcinoma (HCC). Plant alkaloid, harmalol, is being used as a therapeutic agent against HCC due to its accessibility and efficacy by apoptosis and inhibiting proliferation of cancer epithelial cells.ResultSeven groups of Swiss albino mice were taken. Different stages of liver tissues and serum from various experimental groups were collected before and after harmalol treatment. The investigation was carried out by enzyme assay, bilirubin level in the blood, DNA, RNA, normal serum protein of liver tissue, and alpha-feto protein estimation of serum. Gross morphological assessment of liver, histological, and different apoptosis markers viz. p53, caspase3, and cytochrome C expression were analyzed by RT-PCR and Western blot.Harmalol (10 mg/kg B.W. per week, I.P.) for 9 weeks showed a significant reduction in hepatocellular foci, nodules, and carcinoma ultimately retaining the normal morphology. It further induces ROS-dependent apoptosis through mitochondrial cytochrome C release that induces p53 by caspase3 activation.ConclusionThe investigation will eventually help to develop more effective chemotherapeutic drugs from the natural source.