Application status and optimization suggestions of tumor organoids and CAR-T cell co-culture models.

Tumor organoids, especially patient-derived organoids (PDOs) exhibit marked similarities in histopathological morphology, genomic alterations, and specific marker expression profiles to those of primary tumour tissues. They are applied in various fields including drug screening, gene editing, and identification of oncogenes. However, CAR-T therapy in the treatment of solid tumours is still at an exploratory stage. Tumour organoids offer unique advantages over other preclinical models commonly used for CAR-T therapy research, which the preservation of the biological characteristics of primary tumour tissue is critical for the study of early-stage solid tumour CAR-T therapies. Although some investigators have used this co-culture model to validate newly targeted CAR-T cells, optimise existing CAR-T cells and explore combination therapy strategies, there is still untapped potential in the co-culture models used today. This review introduces the current status of the application of tumour organoid and CAR-T cell co-culture models in recent years and commented on the limitations of the current co-cultivation model. Meanwhile, we compared the tumour organoid model with two pre-clinical models commonly used in CAR-T therapy research. Eventually, combined with the new progress of organoid technologies, optimization suggestions were proposed for the co-culture model from five perspectives: preserving or reconstructing the tumor microenvironment, systematization, vascularization, standardized culture procedures, and expanding the tumor organoids resource library, aimed at assisting related researchers to better utilize co-culture models.

Two pairs of phenylpropanoid enantiomers from the leaves of Eucommia ulmoides.

Two pairs of phenylpropanoid enantiomers, (+)-(7S,8S)-alatusol D (1a), (-)-(7R,8R)-alatusol D (1b), (-)-(7S,8R)-alatusol D (2a) and (+)-(7R,8S)-alatusol D (2b) were isolated from the leaves of Eucommia ulmoides Oliver. Among them, 1a and 2b were firstly obtained by chiral enantiomeric resolution. Their structures were elucidated based on extensive spectroscopic analysis and the induced CD (ICD) spectrum caused by adding Mo2(AcO)4 in DMSO. All compounds were tested on Hep G2 tumor cell lines. However, none of the compounds showed potential cytotoxic activity against Hep G2 in vitro.

Zuotai and HgS differ from HgCl2 and methyl mercury in Hg accumulation and toxicity in weanling and aged rats.

Mercury sulfides are used in Ayurvedic medicines, Tibetan medicines, and Chinese medicines for thousands of years and are still used today. Cinnabar (α-HgS) and metacinnabar (ß-HgS) are different from mercury chloride (HgCl2) and methylmercury (MeHg) in their disposition and toxicity. Whether such scenario applies to weanling and aged animals is not known. To address this question, weanling (21d) and aged (450d) rats were orally given Zuotai (54% ß-HgS, 30mg/kg), HgS (α-HgS, 30mg/kg), HgCl2 (34.6mg/kg), or MeHg (MeHgCl, 3.2mg/kg) for 7days. Accumulation of Hg in kidney and liver, and the toxicity-sensitive gene expressions were examined. Animal body weight gain was decreased by HgCl2 and to a lesser extent by MeHg, but unaltered after Zuotai and HgS. HgCl2 and MeHg produced dramatic tissue Hg accumulation, increased kidney (kim-1 and Ngal) and liver (Ho-1) injury-sensitive gene expressions, but such changes are absent or mild after Zuotai and HgS. Aged rats were more susceptible than weanling rats to Hg toxicity. To examine roles of transporters in Hg accumulation, transporter gene expressions were examined. The expression of renal uptake transporters Oat1, Oct2, and Oatp4c1 and hepatic Oatp2 was decreased, while the expression of renal efflux transporter Mrp2, Mrp4 and Mdr1b was increased following HgCl2 and MeHg, but unaffected by Zuotai and HgS. Thus, Zuotai and HgS differ from HgCl2 and MeHg in producing tissue Hg accumulation and toxicity, and aged rats are more susceptible than weanling rats. Transporter expression could be adaptive means to reduce tissue Hg burden.

The Tibetan medicine Zuotai differs from HgCl2 and MeHg in producing liver injury in mice.

Zuotai is composed mainly of ß-HgS, while cinnabar mainly contains α-HgS. Both forms of HgS are used in traditional medicines and their safety is of concern. This study aimed to compare the hepatotoxicity potential of Zuotai and α-HgS with mercury chloride (HgCl2) and methylmercury (MeHg) in mice. Mice were orally administrated with Zuotai (30 mg/kg), α-HgS (HgS, 30 mg/kg), HgCl2 (33.6 mg/kg), or CH3HgCl (3.1 mg/kg) for 7 days, and liver injury and gene expressions related to toxicity, inflammation and Nrf2 were examined. Animal body weights were decreased by HgCl2 and to a less extent by MeHg. HgCl2 and MeHg produced spotted hepatocyte swelling and inflammation, while such lesions are mild in Zuotai and HgS-treated mice. Liver Hg contents reached 45-70 ng/mg in HgCl2 and MeHg groups; but only 1-2 ng/mg in Zuotai and HgS groups. HgCl2 and MeHg increased the expression of liver injury biomarker genes metallothionein-1 (MT-1) and heme oxygenase-1 (HO-1); the inflammation biomarkers early growth response gene (Egr1), glutathione S-transferase (Gst-mu), chemokine (mKC) and microphage inflammatory protein (MIP-2), while these changes were insignificant in Zuotai and HgS groups. However, all mercury compounds were able to increase the Nrf2 pathway genes NAD(P)H: quinone oxidoreductase 1 (Nqo1) and Glutamate-cysteine ligase, catalytic subunit (Gclc). In conclusion, the Tibetan medicine Zuotai and HgS are less hepatotoxic than HgCl2 and MeHg, and differ from HgCl2 and MeHg in hepatic Hg accumulation and toxicological responses.

Simultaneous removal of sulfur dioxide and polycyclic aromatic hydrocarbons from incineration flue gas using activated carbon fibers.

Incineration flue gas contains polycyclic aromatic hydrocarbons (PAHs) and sulfur dioxide (SO2). The effects of SO2 concentration (0, 350, 750, and 1000 ppm), reaction temperature (160, 200, and 280 degrees C), and the type of activated carbon fibers (ACFs) on the removal of SO2 and PAHs by ACFs were examined in this study. A fluidized bed incinerator was used to simulate practical incineration flue gas. It was found that the presence of SO2 in the incineration flue gas could drastically decrease removal of PAHs because of competitive adsorption. The effect of rise in the reaction temperature from 160 to 280 degrees C on removal of PAHs was greater than that on SO2 removal at an SO2 concentration of 750 ppm. Among the three ACFs studied, ACF-B, with the highest microporous volume, highest O content, and the tightest structure, was the best adsorbent for removing SO2 and PAHs when these gases coexisted in the incineration flue gas. Implications: Simultaneous adsorption of sulfur dioxide (SO2) and polycyclic aromatic hydrocarbons (PAHs) emitted from incineration flue gas onto activated carbon fibers (ACFs) meant to devise a new technique showed that the presence of SO2 in the incineration flue gas leads to a drastic decrease in removal of PAHs because of competitive adsorption. Reaction temperature had a greater influence on PAHs removal than on SO2 removal. ACF-B, with the highest microporous volume, highest O content, and tightest structure among the three studied ACFs, was found to be the best adsorbent for removing SO2 and PAHs.

Effects of activated carbon fibre-supported metal oxide characteristics on toluene removal.

Few studies have investigated the use of activated carbon fibres (ACFs) impregnated with metal oxides for the catalytic oxidation of volatile organic compounds (VOCs). Thus, the effects of the ACF-supported metal oxides on toluene removal are determined in this study. Three catalysts, namely, Ce, Mn, and Cu, two pretreatment solutions NaOH and H2O2, and three reaction temperatures of 250 degrees C, 300 degrees C, and 350 degrees C, were employed to determine toluene removal. The composition and morphology of the catalysts were analysed using Brunauer-Emmett-Teller (BET), transmission electron microscope (TEM), inductively coupled plasma (ICP), X-ray photoelectron spectroscopy (XPS), Fourier-transform infrared spectrometer (FTIR), and thermo-gravimetric analyser (TGA) to study the effects of the catalyst's characteristics on toluene removal. The results demonstrated that the metal catalysts supported on the ACFs could significantly increase toluene removal. The Mn/ACFs and Cu/ACFs were observed to be most active in toluene removal at a reaction temperature of 250 degrees C with 10% oxygen content. Moreover, the data also indicated that toluene removal was slightly improved after pretreating the ACFs with NaOH and H2O2. The results suggested that surface-metal loading and the surface characteristics of the ACFs were the determinant parameters for toluene removal. Furthermore, the removal of toluene over Mn/ACFs-H202 decreased when the reaction temperature considered was > 300 degrees C.

Lei's formula attenuates osteoarthritis mediated by suppression of chondrocyte senescence via the mTOR axis: in vitro and in vivo experiments.

Lei's formula (LSF), a traditional Chinese herbal remedy, is recognized for its remarkable clinical effectiveness in treating osteoarthritis (OA). Despite its therapeutic potential, the exact molecular mechanisms underlying LSF's action in OA have remained enigmatic. Existing research has shed light on the role of the mTOR signaling pathway in promoting chondrocyte senescence, a central factor in OA-related cartilage degeneration. Consequently, targeting mTOR to mitigate chondrocyte senescence presents a promising avenue for OA treatment. The primary objective of this study is to establish LSF's chondroprotective potential and confirm its anti-osteoarthritic efficacy through mTOR inhibition. In vivo assessments using an OA mouse model reveal substantial articular cartilage degeneration. However, LSF serves as an effective guardian of articular cartilage, evidenced by reduced subchondral osteosclerosis, increased cartilage thickness, improved surface smoothness, decreased OARSI scores, elevated expression of cartilage anabolic markers (Col2 and Aggrecan), reduced expression of catabolic markers (Adamts5 and MMP13), increased expression of the chondrocyte hypertrophy marker (Col10), and decreased expression of chondrocyte senescence markers (P16 and P21). In vitro findings demonstrate that LSF shields chondrocytes from H2O2-induced apoptosis, inhibits senescence, enhances chondrocyte differentiation, promotes the synthesis of type II collagen and proteoglycans, and reduces cartilage degradation. Mechanistically, LSF suppresses chondrocyte senescence through the mTOR axis, orchestrating the equilibrium between chondrocyte anabolism and catabolism, ultimately leading to reduced apoptosis and decelerated OA cartilage degradation. LSF holds significant promise as a therapeutic approach for OA treatment, offering new insights into potential treatments for this prevalent age-related condition.

[Clinical significance of bronchial reversibility test in the treatment of childhood asthma].

OBJECTIVE: To observe changes in the lung function of asthmatic children with different symptoms during treatment, and to investigate the clinical significance of bronchial reversibility test in the treatment of asthma in children. METHODS: A total of 417 asthmatic children were treated by salmeterol/fluticasone inhalation for more than 3 months. These patients were divided into asymptomatic, single cough, paroxysmal cough and wheeze (cough+wheeze or wheeze alone) groups based on the symptoms when they revisited the clinic. Thirty-four healthy children were used as a control group. All children underwent bronchial reversibility test using nebulized salbutamol. Lung function testing was performed before and after the test. RESULTS: After nebulization of salbutamol, each asthma group showed significantly decreased rate of abnormal lung function and significantly increased forced expiratory volume in one second percent (FEV1%) predicted (P<0.05). Before salbutamol nebulization, the single cough, paroxysmal cough and wheeze groups had significantly higher rates of abnormal lung function and significantly lower FEV1% predicted than the control group (P<0.05). There were significant differences in the rate of abnormal lung function and FEV1% predicted among the asthma groups (P<0.05). After salbutamol nebulization, the paroxysmal cough and wheeze groups had significantly higher rates of abnormal lung function than the control group (P<0.05), but there were no significant differences between other asthma and control groups; the wheeze group had significantly lower FEV1% predicted than the control group, but no significant differences were found between other asthma and the control groups. The positive rate of bronchial reversibility test in each asthma group was significantly higher than in the control group (P<0.05). There were significant differences in the positive rate of the test between the asthma groups except between the asymptomatic and single cough groups (P<0.05). CONCLUSIONS: Asthmatic children with different symptoms demonstrate different lung functions during treatment. Bronchial reversibility test combined with lung function test is useful in assessing asthma control and guiding treatment.

Reference values of gait parameters in healthy Chinese university students: A cross-sectional observational study.

BACKGROUND: Gait is influenced by race, age, and diseases type. Reference values for gait are closely related to numerous health outcomes. To gain a comprehensive understanding of gait patterns, particularly in relation to race-related pathologies and disorders, it is crucial to establish reference values for gait in daily life considering sex and age. Therefore, our objective was to present sex and age-based reference values for gait in daily life, providing a valuable foundation for further research and clinical applications. AIM: To establish reference values for lower extremity joint kinematics and kinetics during gait in asymptomatic adult women and men. METHODS: Spatiotemporal, kinematics and kinetics parameters were measured in 171 healthy adults (70 males and 101 females) using the computer-aided soft tissue foot model. Full curve statistical parametric mapping was performed using independent and paired-samples t-tests. RESULTS: Compared with females, males required more time (cycle time, double-limb support time, stance time, swing time, and stride time), and the differences were statistically significant. In addition, the step and stride lengths of males were longer. Compared to males, female cadence was faster, and statures-per-second and stride-per-minute were higher. There were no statistical differences in speed and stride width between the two groups. After adjusting for height, it was observed that women walked significantly faster than men, and they also had a higher cadence. However, in terms of step length, stride length, and stride width, both genders exhibited similarities. CONCLUSION: We established reference values for gait speed and spatiotemporal gait parameters in Chinese university students. This contributes to a valuable database for gait assessment and evaluation of preventive or rehabilitative programs.

Investigation of the relationship between contact lens design parameters and refractive changes in Ortho-K.

Purpose: To investigate the relationship between Ortho-K contact lens design parameters and refractive power change of the eye through a parametric mathematical representation. Methods: The current study utilises fully anonymized records of 249 eyes, 132 right eyes, and 117 left eyes from subjects aged 14.1 ± 4.0 years on average (range 9-38 years) which were selected for secondary analysis processing. The data were split into 3 groups (G1 up to 35 days wear, from 10 to 35 days, G2 up to 99 days wear, more than 35-99 days & G3 more than 100 days wear) according to the length of time, in days, that the lenses were worn. Corneal shape was measured before and after contact lens wear using the Medmont E300 topographer, from which height and distance files were read by a custom-built MATLAB code to construct the corneal anterior surface independently. Changes in refractive power pre and post-Ortho-K wear were determined using constructed tangential refractive power maps from which both centrally flattened and annular steepened zones were automatically bounded, hence used to determine the refractive power change. Results: On average, flat Sim-K and steep Sim-K were reduced after Ortho-K lens wear by 1.6 ± 1.3 D and 1.3 ± 1.4 D respectively. The radius of the base curve was correlated with the mean central flattened zone power change strongly in G1 (R = 0.7, p < 0.001) and moderately in G2 (R = 0.4) and G3 (R = 0.4, p < 0.001). Hence, a strong correlation with the base curve was recorded in group G1 and moderate in G2 and G3. The reverse curve was very strongly correlated to the mean central flattened zone power change in G1 (R = 0.8, p < 0.001) and strongly correlated with G2 (R = 0.6, p < 0.001) and G3 (R = 0.7, p < 0.001). The reverse curve was also strongly correlated with the mean annular steepened zone power change among all groups G1, G2, and G3 (R = 0.7, R = 0.6 and R = 0.6) respectively (p < 0.001). Conclusions: Although the central corneal refractive power change was strongly correlated to the Ortho-K lens base curve, it characterized only 50% of the target power change. However, the annular steepened zone refractive power change appears to be a clearer predictor of target power change, as there appears to be a one-to-one inverse relationship with the target refractive power correction. Differences between these results and the literature may be a result of the topography software smoothing effect.

[Effects of all-trans retinoic acid on airway responsiveness and airway remodeling in rats with asthma].

OBJECTIVE: To study the effects of alltrans retinoic acid (ATRA) on airway responsiveness, airway remodeling and expression of matrix metalloproteinase-9 (MMP-9) protein in rats with asthma. METHODS: Forty rats were randomly divided into five groups: asthma model, normal saline (control), ATRA treatment, cotton oil treatment and budesonide treatment (n=8 each). Asthma was induced by ovalbumin sensitization and challenge in the asthma model, and the ATRA, cotton oil or budesonide treatment groups. ATRA (50 µg/kg), cotton oil (1 mL) or budesonide (0.32 mg/kg) was administered before ovalbumin challenge in the three treatment groups. Airway responsiveness was assessed. The lung tissues were sampled to detect airway remodeling and the expression of MMP-9 protein by immunohistochemistry. RESULTS: The expression of MMP-9 in lung tissues in the ATRA treatment group was significantly higher than that in the control group, but the airway responsiveness in the ATRA treatment group was not significantly different from that in the control group. The airway responsiveness and the expression of MMP-9 in lung tissues were significantly reduced in the ATRA treatment group compared with the asthma model group. The airway remodeling was significantly improved in the ATRA treatment group compared with the asthma model group. CONCLUSIONS: ATRA may alleviate airway hyperresponsiveness and airway remodeling possibly through decreasing the protein expression of MMP-9 in rats with asthma.

Yinchenzhufu decoction protects against alpha-naphthylisothiocyanate-induced acute cholestatic liver injury in mice by ameliorating disordered bile acid homeostasis and inhibiting inflammatory responses.

ETHNOPHARMACOLOGICAL RELEVANCE: Intrahepatic cholestasis is a common condition of many liver diseases with few therapies. Yinchenzhufu decoction (YCZFD) is a representative traditional Chinese herbal formula used for treating jaundice and liver disease. AIM OF THE STUDY: To investigate the hepatoprotective effect of YCZFD against cholestatic liver injury and reveal its potential mechanism. MATERIALS AND METHODS: Mice with alpha-naphthyl isothiocyanate (ANIT)-induced intrahepatic cholestasis were orally administered YCZFD at doses of 3, 6, and 12g crude drug/kg for 2 weeks followed by subsequent analyses. A serum metabolomics study was then performed to explore the different metabolites influenced by YCZFD using ultra-high-performance liquid chromatography coupled with linear ion trap-Orbitrap hybrid mass spectrometry (UPLC-LTQ-Orbitrap-MS/MS).The levels of individual bile acids in the serum, liver, and bile were determined by UPLC-MS/MS. The expression of metabolic enzymes, transporters, inflammatory factors, and cytokeratin-19 (CK-19) was determined by real-time PCR, western blotting, and immunohistochemistry. RESULTS: YCZFD administration decreased the serum biochemical indexes and ameliorated pathological damage, such as hepatic necrosis and inflammatory cell infiltration. Serum metabolomics revealed that the metabolites influenced by YCZFD were mainly associated with bile acid metabolism and inflammation. YCZFD administration effectively ameliorated the disordered bile acid homeostasis. The bile acid transporter, multidrug-resistance associated protein 2 (Mrp2), and the metabolic enzyme, cytochrome P450 2b10 (Cyp2b10), were upregulated in the YCZFD intervention group compared to those in the ANIT-induced group. YCZFD administration also significantly inhibited nuclear factor-κB (NF-κB) and its phosphorylation and decreased the expression of proinflammatory cytokines including tumor necrosis factor-α, interleukin-1ß, and intercellular adhesion molecule-1 in ANIT-induced cholestatic mice. Additionally, the level of CK-19 was lower in the YCZFD intervention group than in the ANIT-induced cholestatic mice. CONCLUSION: YCZFD administration ameliorated disordered bile acid homeostasis, inhibited NF-κB pathway-mediated inflammation, and protected the liver from bile duct injury. Therefore, YCZFD exerted a protective effect against cholestatic liver injury.

Protective effect of herbal medicine Huangqi decoction against chronic cholestatic liver injury by inhibiting bile acid-stimulated inflammation in DDC-induced mice.

BACKGROUND: Huangqi decoction (HQD), a classic traditional herbal medicine, has been used for liver fibrosis, but its effect on intrahepatic chronic cholestatic liver injury remains unknown. PURPOSE: In the present study, we investigated the hepatoprotective effect of HQD and the underlying molecular mechanisms in 3, 5-diethoxycarbonyl-1, 4-dihydroxychollidine (DDC)-induced chronic cholestatic mice. METHODS: The DDC-induced cholestatic mice were administrated HQD for 4 or 8 weeks. Serum biochemistry and morphology were investigated. The serum and liver bile acid (BA) levels were detected by ultra performance liquid chromatography-tandem mass spectrometry. The liver expression of BA metabolizing enzymes and transporters, and inflammatory and fibrotic markers was measured by real-time polymerase chain reaction, western blotting, and immunohistochemistry. RESULTS: HQD treatment for 4 or 8 weeks ameliorated DDC-induced liver injury by improving impaired hepatic function and tissue damage. HQD treatment for 8 weeks further decreased the liver expression of cytokeratin 19, tumor growth factor (TGF)-ß, collagen I, and α-smooth muscle actin, and ameliorated ductular reaction and liver fibrosis. HQD markedly decreased the accumulation of serum and liver BA. The expression of BA-metabolizing enzymes, cytochrome P450 2b10 and UDP glucuronosyltransferase 1 A1, and multidrug resistance-associated protein 2, Mrp3, and Mrp4 involved in BA homeostasis was increased by 4 weeks of HQD treatment. The expression of BA uptake transporter Na+-taurocholate cotransporting polypeptide was decreased and that of Mrp4 was increased after 8 weeks of HQD treatment. Nuclear factor-E2-related factor-2 (Nrf2) was remarkably induced by HQD treatment. Additionally, HQD treatment for 8 weeks decreased the liver expression of inflammatory factors, interleukin (IL)-6, IL-1ß, tumor necrosis factor-α, monocyte chemoattractant protein-1, and intracellular adhesion molecule-1. HQD suppressed the nuclear factor (NF)-κB pathway. CONCLUSION: HQD protected mice against chronic cholestatic liver injury and biliary fibrosis, which may be associated with the induction of the Nrf2 pathway and inhibition of the NF-κB pathway, ameliorating BA-stimulated inflammation.

[Sex determination by discriminant analysis of calcaneal measurements on the lateral digital radiography].

OBJECTIVE: Five measurements of the calcaneus taken on digital radiography (DR) of adults of Han Population of Sichuan Province were selected to determine sex by multivariate discriminant analysis. METHODS: Lateral radiographs of calcaneus taken from 393 subjects were collected. The samples were randomly divided into the experimental group (148 males and 186 females) and the examined group (26 males and 33 females). Five measurements were taken from the radiography. The analysis of variance (AVON) was carried out to determine if there was significant difference between the male and female. The discriminant functions were drawn by Fisher discriminant analysis. The effects of all obtained functions were evaluated with the examined samples. RESULTS: There was statistically significant difference in the five measurements between the males and the females (P<0.05). Six groups of discriminant functions were obtained with an accuracy ranged from 78.4% to 88.9%. When applied on the examined samples, the sex discriminant accuracy varied from 79.7% to 86.4%. CONCLUSION: These five measurements acquired from the lateral radiographs of calcaneus could be used for sex assessment during forensic identification of individuals.

[Effects of ginsenoside Rg1 on expression of insulin-like growth factor-1 in brain of rats with brain contusion].

OBJECTIVE: To study the effects of ginsenoside Rg1 on the expression of insulin-like growth factor-1 (IGF-1) in the brain of rats after the experimental brain contusion. METHODS: A total of twenty-six Wistar rats were randomly divided into normal control group (n=2), untreated group (n=8) and ginsenoside Rg1 group (n=16). Brain injuries were induced in rats by a mechanical striking device. The brain tissues were extracted at different times after brain injury (6th hour, 12th hour, 2nd day, 6th day), then the expression of IGF-1 in brain tissue was examined by immunohistochemical method. RESULTS: In comparison with the normal control group, the expression of IGF-1 in the brain tissues was increased in the untreated group after the brain contusion (P<0.05). The expression of IGF-1 in brain tissues in ginsenoside Rg1 group was significantly increased as compared with the untreated group (P<0.05). CONCLUSION: Ginsenoside Rg1 enhances the recovery of the contused brain through increasing the expression of IGF-1.

Atorvastatin alters the expression of genes related to bile acid metabolism and circadian clock in livers of mice.

AIM: Atorvastatin is a HMG-CoA reductase inhibitor used for hyperlipidemia. Atorvastatin is generally safe but may induce cholestasis. The present study aimed to examine the effects of atorvastatin on hepatic gene expression related to bile acid metabolism and homeostasis, as well as the expression of circadian clock genes in livers of mice. METHODS: Adult male mice were given atorvastatin (10, 30, and 100 mg/kg, po) daily for 30 days, and blood biochemistry, histopathology, and gene expression were examined. RESULTS: Repeated administration of atorvastatin did not affect animal body weight gain or liver weights. Serum enzyme activities were in the normal range. Histologically, the high dose of atorvastatin produced scattered swollen hepatocytes, foci of feathery-like degeneration, together with increased expression of Egr-1 and metallothionein-1. Atorvastatin increased the expression of Cyp7a1 in the liver, along with FXR and SHP. In contract, atorvastatin decreased the expression of bile acid transporters Ntcp, Bsep, Ostα, and Ostß. The most dramatic change was the 30-fold induction of Cyp7a1. Because Cyp7a1 is a circadian clock-controlled gene, we further examined the effect of atorvastatin on clock gene expression. Atorvastatin increased the expression of clock core master genes Bmal1 and Npas2, decreased the expression of clock feedback genes Per2, Per3, and the clock targeted genes Dbp and Tef, whereas it had no effect on Cry1 and Nr1d1 expression. CONCLUSION: Repeated administration of atorvastatin affects bile acid metabolism and markedly increases the expression of the bile acid synthesis rate-limiting enzyme gene Cyp7a1, together with alterations in the expression of circadian clock genes.

Huangqi Decoction Alleviates Alpha-Naphthylisothiocyanate Induced Intrahepatic Cholestasis by Reversing Disordered Bile Acid and Glutathione Homeostasis in Mice.

Intrahepatic cholestasis is a serious symptom of liver disorders with limited therapies. In this study, we investigated the efficacy of Huangqi decoction (HQD), a two-herb classic traditional Chinese medicine (TCM), in the treatment of alpha-naphthylisothiocyanate (ANIT)-induced intrahepatic cholestasis in mice. HQD treatment ameliorated impaired hepatic function and tissue damage. A metabolomics study revealed that the endogenous metabolites significantly affected by HQD were related to bile acid (BA) biosynthesis and glutathione metabolism pathways. HQD treatment decreased the intrahepatic accumulation of cytotoxic BAs, normalized serum BA levels, and increased biliary and urinary BA excretion. Additionally, HQD restored the hepatic glutathione content and suppressed reactive oxygen species (ROS) in cholestatic mice. Protein and gene analysis revealed that HQD increased the expression of the hepatic metabolizing enzymes cytochrome P450 (CYP) 2B10 and UDP glucuronosyltransferase family 1 member A1 (UGT1A1), as well as multidrug resistance-associated protein 2 (Mrp2), Mrp3, and Mrp4, which play crucial roles in BA homeostasis. Further, HQD increased the protein expression of glutamate-cysteine ligase, which is involved in the synthesis of glutathione. Importantly, HQD increased the nuclear expression of nuclear factor-E2-related factor-2 (Nrf2). In conclusion, HQD protects against intrahepatic cholestasis by reversing the disordered homeostasis of BAs and glutathione.

Chicken bile powder protects against α-naphthylisothiocyanate-induced cholestatic liver injury in mice.

This study explored the effects of chicken bile powder (CBP), a 2000-year-old Chinese medicine, on α-naphthyl isothiocyanate (ANIT)-induced intrahepatic cholestasis in mice. CBP treatment for 14 days significantly ameliorated ANIT-induced changes in serum alanine aminotransferase, aspartate aminotransferase, bile acids, bilirubin, γ-glutamyl transpeptidase, alkaline phosphatase, and liver tissue morphology. Serum metabolomics showed changes in 24 metabolites in ANIT-exposed mice; 16 of these metabolites were reversed by CBP treatment via two main pathways (bile acid biosynthesis and arachidonic acid metabolism). Additionally, CBP administration markedly increased fecal and biliary bile acid excretion, and reduced total and hydrophobic bile acid levels in the livers of cholestatic mice. Moreover, CBP increased liver expression of bile acid efflux transporters and metabolic enzymes. It also attenuated ANIT-induced increases in hepatic nuclear factor-κB-mediated inflammatory signaling, and increased liver expression of the nuclear farnesoid X receptor (FXR) in cholestatic mice. CBP also activated FXR in vitro in HEK293T cells expressing mouse Na+-taurocholate cotransporting polypeptide. It did not ameliorate the ANIT-induced liver injuries in FXR-knockout mice. These results suggested that CBP provided protection from cholestatic liver injury by restoring bile acid homeostasis and reducing inflammation in a FXR-dependent manner.

[Cloning and expression of the succinate dehydrogenase iron-sulfur protein of Schistosoma japonicum Chinese strain in E. coli].

OBJECTIVE: To clone the full-length gene encoding succinate dehydrogenase iron-sulfur protein of Schistosoma japonicum (SjSDISP) Chinese strain and express it in Escherichia coli. METHODS: According to the published incomplete EST (BU804141) of SjSDISP and the sequence of multiclone sites of lambda gt11 vector, 2 pairs of primers were designed and synthesized. Then the 3' and 5'ends of the EST of the SjSDISP from adult Schistosoma japonicum cDNA library were amplified by anchored PCR. After sequencing, a full-length cDNA sequence of the SjSDISP was obtained, and then it was cloned into prokaryotic expression vector pGEX-4T-1. Identified by agarosed gel electrophoresis, endonucleases digestion and PCR, the resultant recombinant plasmid was used for the expression under the temperature-dependent condition and Western blot analysis. RESULTS: A 1,071 bp sequence was obtained. Sequence analysis showed that the fragment contained a complete open reading frame (ORF), encoding 278 amino acid residues. This target fragment was cloned into the prokaryotic expression vector pGEX-4T-1, and expressed in Escherichia coli. SDS-PAGE revealed that the molecular weight of the expressed fusion recombinant product was 56 kD. Western blot showed that the recombinant protein was recognized by polyclonal rabbit antiserum immunized with Schistosoma japonicum adult worm antigen. CONCLUSION: Cloning of the full-length gene encoding SjSDISP and its bacterial expression were successfully done.

The Tibetan medicine Zuotai influences clock gene expression in the liver of mice.

Background. The circadian clock is involved in drug metabolism, efficacy and toxicity. Drugs could in turn affect the biological clock as a mechanism of their actions. Zuotai is an essential component of many popular Tibetan medicines for sedation, tranquil and "detoxification," and is mainly composed of metacinnabar (ß-HgS). The pharmacological and/or toxicological basis of its action is unknown. This study aimed to examine the effect of Zuotai on biological clock gene expression in the liver of mice. Materials and methods. Mice were orally given Zuotai (10 mg/kg, 1.5-fold of clinical dose) daily for 7 days, and livers were collected every 4 h during the 24 h period. Total RNA was extracted and subjected to real-time RT-PCR analysis of circadian clock gene expression. Results. Zuotai decreased the oscillation amplitude of the clock core gene Clock, neuronal PAS domain protein 2 (Npas2), Brain and muscle Arnt-like protein-1 (Bmal1) at 10:00. For the clock feedback negative control genes, Zuotai had no effect on the oscillation of the clock gene Cryptochrome (Cry1) and Period genes (Per1-3). For the clock-driven target genes, Zuotai increased the oscillation amplitude of the PAR-bZip family member D-box-binding protein (Dbp), decreased nuclear factor interleukin 3 (Nfil3) at 10:00, but had no effect on thyrotroph embryonic factor (Tef); Zuotai increased the expression of nuclear receptor Rev-Erbα (Nr1d1) at 18:00, but had little influence on the nuclear receptor Rev-Erbß (Nr1d2) and RORα. Conclusion. The Tibetan medicine Zuotai could influence the expression of clock genes, which could contribute to pharmacological and/or toxicological effects of Zuotai.