A PETAR method for risk assessment of human health and environment on the regional scale.

Risk assessments are necessary to effectively reveal the state of the degradation of living environments on a regional scale. However, risk assessments are often limited by time, cost, and technology, which make conducting effective evaluations difficult. Thus, in this study, the procedure for ecological tiered assessment of risk (PETAR) method was used to analyze the human health and environmental risks in Daye, China. This method first used the United States Environmental Protection Agency's risk assessment approach to qualitatively determine the risk sources, pressures, receptors, and effect endpoints and constructed a conceptual model of threats to the human living environment. Each risk-prone subregion was then evaluated using the fuzzy logic method. Next, a quantitative assessment was conducted for the subregions with the most serious environmental degradation. Finally, quantitative analyses were performed to verify the original hypotheses. The results showed that the high-risk areas were distributed in the industrial regions of Daye, wherein mining and processing clusters and mining settlements are widespread and confirmed the locations of the particular subregions with the most serious human health and environmental risks. This study also validated the practicality of the PETAR method for human health risk assessments in mining areas with large-scale, multifactor, and multihazard paths. Integr Environ Assess Manag 2023;19:239-253. © 2022 SETAC.

Rapid Screening and Characterization of Triterpene Saponins from the Root of Phytolacca acinosa Roxb by High-Performance Liquid Chromatography Coupled to Electrospray Ionization and Quadrupole Time-of-Flight Mass Spectrometry.

Triterpene saponins (TSs) are important bioactive constituents with structural diversity widely distributed in many plants. The root of Phytolacca acinosa Roxb (RPa) has been used as a traditional Chinese medicine. However, TSs as the main active ingredients in RPa have not been fully characterized. Here, we profiled TSs from RPa by high-performance liquid chromatography coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry (HPLC-ESI-QTOF-MS/MS). We tentatively identified 29 TSs, including 13 that had not been reported previously from this plant. This study indicates that HPLC-ESI-QTOF-MS/MS is an effective and rapid method for the characterization of complicated TSs in herbal extracts.

Protopine inhibits the growth of hepatocellular carcinoma through a mitochondrially mediated apoptosis pathway / 药学学报

Hepatocellular carcinoma (HCC) is a serious threat for human health, the incidence of HCC in China accounts for more than 50% worldwide. There is an urgent need to develop novel anticancer agents for the treatment of HCC patients. Here we characterized the inhibitory effect and the molecular mechanism of protopine on HCC cancer cells. The results of a CCK-8 assay indicated that protopine displays anticancer activities on HCC cells. Flow cytometry and JC-1 staining confirmed that treatment with protopine decreased the mitochondrial membrane potential and induced apoptosis in HCC cells. Western blot analysis showed that protopine was able to increase protein expression in the mitochondrial apoptotic pathway; the level of cytochrome C, apoptotic protease activating factor-1 (Apaf-1), Bax, cleaved-poly ADP-ribose polymerase (cleaved-PARP), cleaved-caspase-3, and cleaved-caspase-9 were increased while the expression of Bcl-2 was suppressed significantly. An in vivo study revealed that protopine significantly suppressed the growth of tumors in nude mice bearing HepG2 cells. Administration of protopine intraperitoneally at a concentration of 50 mg·kg-1 inhibited tumor growth by 72.46%. Animal experiments were carried out according to the Regulation of the Animal Ethics Committee of Southwest Medical University. This study provides preliminary evidence that there is potential to develop protopine as a lead compound for the treatment of HCC.

Preparation of exosome-coated Bcl-2 siRNA and its inhibitory effect on the growth of pancreatic cancer cells / 药学学报

High expression of Bcl-2 is associated with the development of pancreatic cancer, and downregulation of Bcl-2 is an effective approach for the treatment of pancreatic malignancy. In the present study exosomes were isolated from the cultured medium of human embryonic kidney cells (HEK293) by ultracentrifugation and exosome-coated Bcl-2 siRNA (exosiBcl-2) was synthesized using electroporation. The results showed that the particle size of exosiBcl-2 was 67.3 ± 9.7 nm and the morphology of exosomes displayed a concave ring structure as determined by transmission electron microscopy (TEM). Western blot analysis indicated that exosomal proteins including CD9, CD81, CD63 and TSG101 were highly expressed. Confocal microscopy revealed that exosiBcl-2 was widely distributed in Miapaca-2 cells, and the transfection efficiency of exosiBcl-2 in Miapaca-2 was 77.2% as determined by flow cytometry. Treatment with exosiBcl-2 at a concentration of 100 nmol·L-1 resulted in an inhibitory effect on the growth of Miapaca-2 cells with an inhibition rate of 63%. ExosiBcl-2 treatment can downregulate Bcl-2 and upregulate Bax protein. This study provides evidence that exosiBcl-2 is able to inhibit the growth of pancreatic cancer cells and the nanoparticles have potential to be developed as a novel anticancer agent.

Highly efficient biocatalytic synthesis of L-DOPA using in situ immobilized Verrucomicrobium spinosum tyrosinase on polyhydroxyalkanoate nano-granules.

L-DOPA (3,4-dihydroxyphenyl-L-alanine) is a preferred drug for Parkinson's disease, and is currently in great demand every year worldwide. Biocatalytic conversion of L-tyrosine by tyrosinases is the most promising method for the low-cost production of L-DOPA in both research and industry. Yet, it has been hampered by low productivity, low conversion rate, and low stability of the biocatalyst, tyrosinase. An alternative tyrosinase TyrVs from Verrucomicrobium spinosum with more efficient expression in heterologous host and better stability than the commercially available Agaricus bisporus tyrosinase was identified in this study. Additionally, it was prepared as a novel nano-biocatalyst based on the distinct one-step in situ immobilization on the surface of polyhydroxyalkanoate (PHA) nano-granules. The resulting PHA-TyrVs nano-granules demonstrated improved L-DOPA-forming monophenolase activity of 9155.88 U/g (Tyr protein), which was 3.19-fold higher than that of free TyrVs. The nano-granules also exhibited remarkable thermo-stability, with an optimal temperature of 50 °C, and maintained more than 70% of the initial activity after incubation at 55 °C for 24 h. And an enhanced affinity of copper ion was observed in the PHA-TyrVs nano-granules, making them even better biocatalysts for L-DOPA production. Therefore, a considerable productivity of L-DOPA, amounting to 148.70 mg/L h, with a conversion rate of L-tyrosine of 90.62% can be achieved by the PHA-TyrVs nano-granules after 3 h of biocatalysis under optimized conditions, without significant loss of enzyme activity or L-DOPA yield after 8 cycles of repeated use. Our study provides an excellent and robust nano-biocatalyst for the cost-effective production of L-DOPA.

Novel Zn-doped CuO nanocomposites inhibit tumor growth in vitro and in vivo:role of reactive oxygen species-dependent apoptosis and autophagy cross-linked by NF-kappaB pathway / 中国药理学与毒理学杂志

OBJECTIVE Zn-doped CuO nanocomposites (nZn-CuO NPs) are novel nanoparticles synthesized by sonochemical method.This study aimed to further investigate the antitumor effects and mechanism of nZn-CuO NPs, as well as the exact mechanism of reactive oxygen species (ROS) on nZn-CuO NPs-induced death using N-acetylcysteine (NAC). METHODS The antitumor effects of nZn-CuO NPs were evaluated by MTS assay and orthotopic transplantation tumor model in nude mice. The effects of nZn-CuO NPs with or without NAC on ROS production, DNA damage, apoptosis, mitochondrial damage, autophagy, lysosome impairment, and ER and Golgi stress were determined. Also,western blot was used to detect apoptosis and autophagy related proteins,as well as NF-κB pathway related proteins. RESULTS nZn-CuO NPs significantly inhibit tumor growth both in vitro and in vivo. nZn-CuO NPs were able to cause cytotoxicity, ROS production, DAN damage mitochondrial damage, apoptosis, and autophagy, and NAC can attenuate them. Further studies showed that nZn-CuO NPs induced changes of apoptosis, autophagy and NF-κB pathway related proteins, and NAC can restore them. CONCLUSION Overall, our data demonstrated that nZn-CuO NPs could inhibit tumor growth both in vitro and in vivo by ROS-dependent regulation of apoptosis and autophagy, which might be cross-linked by NF-κB pathways.

Granulin A synergizes with cisplatin to inhibit the growth of human hepatocellular carcinoma / 中国药理学与毒理学杂志

OBJECTIVE Granulin A (GRN A), a cytokinesis protein, is derived from proteolysis of progranulin. The previous study in our laboratory has shown that GRN A is able to inhibit cancer cell growth significantly. This study aimed to investigate the effect of combination of GRN A and cisplatin on in vitro and in vivo on the growth of hepatocellular carcinoma. METHODS The in vitro and in vivo antitumor effects of combination of GRN A and Cisplatin were evaluated with MTS assay and subcuta-neous transplantation tumor model.Chou-Talalay method was used to calculate the combination index (CI). Colony formation assay and flow cytometry were used to detect the effects of GRN A on apoptosis. The expression of apoptosis-related proteins were detected by Western blot. RESULTS MTS assay showed that GRN A significantly inhibit hepatocellular carcinoma cells growth with the IC50of 5.6 μmol·L-1, and GRN A combined with cisplatin synergistically inhibit hepatocellular carcinoma proliferation, with the CI<1.The colony-formation assay showed that GRN A significantly enhanced the inhibitory effects of cisplatin on cellular anchorage-independent growth. Flow cytometry showed that GRN A combined with cisplatin synergistically induced apoptosis,with the apoptotic rates of 5.87%,32.74%,35.67% and 67.15% in control, GRN A, Cisplatin, and combination of GRN A and Cisplatin groups, respectively. Western blot confirmed that the two drugs synergistically changed the expressions of proteins related to apoptosis.In vivo experiment indicated that combination of GRN A and cisplatin significantly suppressed tumor growth compared with single drug treatment groups.CONCLUSION The combination of GRN A and cisplatin resulted in synergistic antitumor effects against hepatocellular carcinoma both in vitro and in vivo.

Zn-doped CuO nanocomposites inhibit in vitro and in vivo growth of pancreatic cancer by inducing autophagy through AMPK/mTOR pathway / 中国药理学与毒理学杂志

OBJECTIVE Zn-doped CuO nanocomposites (Zn-CuO NPs) are novel nanoparticles synthesized by our research group.In this study,we assessed the in vitro and in vivo antitumor effects of Zn-CuO NPS on pancreatic cancer cells,as well as the potential mechanisms. METHODS MTS assay was used to detect the effects of Zn-CuO NPS on proliferation pancreatic cancer cells(Panc-mia and Aspc-1). The in vivo antitumor effects of Zn-CuO NPs were detected by xenografts model in nude mice. The effects of Zn-CuO NPS on autophagy were detected bytransmission electron microscopy (TEM) andflow cytometry. Autophagy related proteins were detected by Western blotting. RESULTS Zn-CuO NPS significantly inhibited the proliferation of Panc-mia cells and Aspc-1 cells.In vivo experi-ments showed that Zn-CuO NPS significantly inhibited the tumor growth in nude mice without affecting the body weight of the mice. TEM and flow cytometry showed that Zn-CuO NPS induced autophagy, and significantly increased the number of autophagosome.Western Blot showed that Zn-CuO NPS alterd the expression of autophagy related proteins,such as AMPK,mTORand Beclin-1.Also,AMPK inhibitor could significantly reduce Zn-CuO NPS-induced autophagy pathwayas analyzed byWestern blotting. CONCLUSION The findings suggested that Zn-doped CuO nanocomposites inhibited the in vitro and in vivo growth of pancreatic cancer by inducing autophagy through AMPK/mTOR pathway.

Water-soluble extract of clove inhibits in vitro and in vivo growth of colon cancer by inducing autophagy / 中国药理学与毒理学杂志

OBJECTIVE Cloves(Syzygium aromaticum L.)have been used as both a spice and a traditional Chinese medicinal herb for thousands of years. However, relatively little is known about its potential anticancer activity and mechanisms.In this study,we investigated the in vitro and in vivo anti-tumor effects and mechanisms of water extract of cloves(WEC)against colorectal cancer. METHODS MTS assay and Colony-formation assay were used to detect the anti-tumor activity of WEC on HT-29 cells.The in vivo anti-tumor effect of WEC was detected in a subcutaneous transplantation tumor model of human HT-29 cells.Autophagy was detected by flow cytometry and the expressions of autophagy related proteins(Beclin-1 and LC-3a/b)were determined by western blot. RESULTS MTS result showed that WEC significantly inhibited the viability of HT-29 cells,with the IC50values of 150 μg·mL-1.The colony-formation assay showed that the WEC significantly suppressed colon cancer cells proliferation.WEC also exhibited significant antitumor activity in tumor bearing nude mice. Flow cytometry result showed that WEC significantly induced autophagy, and the averaged relative values of fluorescence intensity were 206,251,341 and 356 in cells treated with 0,100,150 and 200 μg·mL-1WEC for 48 h.Western blot result showed that WEC treatment significantly increased Beclin-1 expression and ratios of LC3-II/LC3-I. CONCLUSION These result showed that WEC inhibited the growth of colon tumor both in vitro and in vivo, which might be related with autophagy induction, and WEC has potential to be developed as a novel anticancer agent for the treatment of colon cancer.

[Risk factors of hepatorenal syndrome in patients with acute on chronic liver failure].

OBJECTIVE: To identify the risk factors of hepatorenal syndrome in patients with hepatitis B virus (HBV)-related acute-on-chronic liver failure(ACLF). METHODS: A total of 726 hospitalized patients with HBV-ACLF were retrospectively analyzed. Data of demographic and clinical parameters (sex, age, family history, and presence of liver cirrhosis and diabetes), common complications (spontaneous bacterial peritonitis, pulmonary infection, hepatic encephalopathy, and upper gastrointestinal hemorrhage), and baseline biochemical parameters (albumin, globulin, total bilirubin, direct bilirubin, alanine aminotransferase, aspartate aminotransferase, gamma-glutamyl transferase, alkaline phosphatase, cholesterol, cholinesterase, K+, Na+, plasma thromboplastin antecedent, alpha-fetoprotein, HBV DNA, white blood cell count, hemoglobin, and platelet count) were collected from the medical records database. Univariate and multiple regression analyses were performed to determine the risk factors of hepatorenal syndrome. RESULTS: Multiple logistic regression analysis indicated that upper gastrointestinal hemorrhage [risk (R) = 1.313, relative hazard (RH) = 3.716, 95% confidence interval (CI): 2.156-6.404], hepatic encephalopathy (R = 1.120, RH = 3.065, 95% CI: 1.900-4.945), spontaneous bacterial peritonitis (R = 1.005, RH = 2.733, 95% CI: 1.379-5.417), pulmonary infection (R = 1.051, RH = 2.862, 95% CI: 1.783-4.592), and white blood cell count (R = 0.056, RH = 1.058, 95% CI: 1.010-1.107) were independent risk factors for hepatorenal syndrome development in patients with HBV-ACLF. CONCLUSION: Several risk factors were significantly associated with the development of hepatorenal syndrome in HBV-ACLF, including upper gastrointestinal hemorrhage, hepatic encephalopathy, spontaneous bacterial peritonitis, pulmonary infection, and elevated white blood cell count.

[Clinical characteristics of and factors related to relapse in chronic hepatitis B patients after nucleoside analogues withdrawal].

OBJECTIVE: To investigate the clinical characteristics of and factors related to relapse in chronic hepatitis B (CHB) patients who had previously achieved cessation criteria and had been withdrawn from nucleoside analogues treatment. METHODS: Sixty CHB patients who experienced relapse after nucleoside analogues withdrawal based on cessation criteria were enrolled in the study retrospectively. Each patient's data on biochemical, serological and viral characteristics corresponding to baseline (treatment initiation), withdrawal and relapse were collected. COX proportional hazard modeling was used to evaluate the factors related to relapse. RESULTS: The hepatitis B e antigen (HBeAg)-positive and -negative patients had similar median antiviral treatment times (38 months (range: 24 - 80) vs. 35 months (30 - 60); Z = -1.313, P more than 0.05). For all patients, the median follow-up time was 12 months (2 - 72), during which 49 (81.7%) patients developed virological breakthrough and 17 (28.3%) developed HBeAg recurrence. The patients who experienced virological breakthrough or HBeAg recurrence had significantly higher baseline levels of HBV DNA than those patients who remained disease-free (t = 2.15 and -2.54 respectively; P less than 0.05). The median relapse time of the HBeAg-positive patients was significantly longer than that of the HBeAg-negative patients (14 months (3 - 72) vs. 6 months (3 - 36); Chi-square test = 7.045, P less than 0.01). HBeAg status at baseline was identified as an independent factor associated with relapse (relative risk = 1.937, 95% confidence interval = 1.14-3.28, P less than 0.05). CONCLUSION: HBeAg-positive and-negative patients showed distinct clinical characteristics of relapse, with the latter being more prone to relapse soon after nucleoside analogues withdrawal. Prolonging the treatment course may be beneficial to HBeAg-negative patients, even if cessation criteria are achieved.

[The impact of early rapid virological response on the outcomes of hepatitis B associated acute on chronic liver failure during antiviral treatment].

OBJECTIVE: To investigate the impact of early rapid virological response on the outcomes of hepatitis B associated acute on chronic liver failure during antiviral treatment. METHODS: 106 acute on chronic liver failure patients in our hospital from January 2008 to July 2010 were enrolled in present study retrospectively. Besides internal medicine therapy, all patients received lamivudine (100 mg/d) or entecavir (0.5 mg/d) treatment. The profile of liver biochemistry, prothrombin time activity and viral load were detected at baseline and week 4, respectively. The patients were divided into HBV DNA negative group and HBV DNA positive group according to the viral load at week 4. The clinical features and treatment outcomes were compared between groups. Frequency variables were compared by x2 test or Fisher's exact test. Continuous variables were compared using independent samples T-test. The factors that impact on the treatment outcomes were determined using stepwise multivariate logistic regression analysis. RESULTS: At the week 4, the TBil and PTA in HBV DNA positive group [(261.6+/-205.6)mumol/L and 44.7%+/-19.7%, respectively] were significantly different from those in HBV DNA negative group [(160.1+/-173.4) mumol/L and 56.8%+/-23.1%, respectively] ( t = 2.190 and -2.077, respectively, P less than 0.05). The non-effective rate of HBVDNA positive group (50%, 9/18) was significantly higher than that of HBV DNA negative group (14.8%, 13/88) (x2 = 9.235, P less than 0.01). By using stepwise multivariate logistic regression analysis, the disease stage and HBV DNA undetectable at week 4 were the independent factor. The OR values of disease stage and HBV DNA undetectable were 6.559 and 0.209, respectively, and 95% CI was 2.316~18.576 and 0.058~0.747, respectively. CONCLUSION: The rapid suppression of viral load by nucleotide analogue may improve the efficacy of hepatitis B associated acute on chronic liver failure treatment. The early rapid virological response within first 4 weeks may contribute to the prediction of the treatment outcomes.