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Sarcomatoid mesothelioma is an aggressive malignancy that can be challenging to distinguish from benign spindle cell mesothelial proliferations based on biopsy, and this distinction is crucial to patient treatment and prognosis. A novel deep learning based classifier may be able to aid pathologists in making this critical diagnostic distinction. SpindleMesoNET was trained on cases of malignant sarcomatoid mesothelioma and benign spindle cell mesothelial proliferations. Performance was assessed through cross-validation on the training set, on an independent set of challenging cases referred for expert opinion (‘referral’ test set), and on an externally stained set from outside institutions (‘externally stained’ test set). SpindleMesoNET predicted the benign or malignant status of cases with AUC’s of 0.932, 0.925, and 0.989 on the cross-validation, referral and external test sets, respectively. The accuracy of SpindleMesoNET on the referral set cases (92.5%) was comparable to the average accuracy of 3 experienced pathologists on the same slide set (91.7%). We conclude that SpindleMesoNET can accurately distinguish sarcomatoid mesothelioma from benign spindle cell mesothelial proliferations. A deep learning system of this type holds potential for future use as an ancillary test in diagnostic pathology.

Nanostructured phenolic resin-based carbon aerogels with an extensive network structure are regarded as ideal energy storage materials for supercapacitors. However, the initial bulk form and low capacitance of previously reported porous carbon aerogels are problematic for practical use. Phenolic resin-based porous carbon spheres were synthesized by a simple hydrothermal process using ammonia, ethylenediamine or hexylenediamine as a catalyst. The porous carbon spheres were investigated by SEM, BET, XPS, etc. It was found that the number of ammonium groups, length of the alkyl chain and processing temperature play vital roles in determining the pore structure, size and uniformity of the carbon spheres. NH4+ is necessary to obtain the carbon spheres and but changing the other parameters has no obvious effect on their crystal structure. The sample prepared at a hydrothermal temperature of 80 °C using ammonia as the catalyst has the highest specific capacitance of 233.8 F g−1 at a current density of 1.0 A g−1. It has an excellent capacitance retention of 98% after 10,000 charge/discharge cycles at 7 A g−1, indicating its good cycling stability and rate capability. This result shows that a higher specific surface area, porosity and defect density are probably the crucial factors in improving the electrochemical capacitance.

Metabotropic glutamate receptor (mGlu)5 regulates microglia activation, which contributes to inflammation. However, the role of mGlu5 in neuroinflammation associated with Parkinson's disease (PD) remains unclear. Triptolide (T10) exerts potent immunosuppressive and anti-inflammatory effects and protects neurons by inhibiting microglia activation. In this study, we investigated the role of mGlu5 in the anti-inflammatory effect of T10 in a lipopolysaccharide (LPS)-induced PD model. In cultured BV2 cells and primary microglia, blocking mGlu5 activity or knocking down its expression abolished T10-inhibited release of proinflammatory cytokines induced by LPS. Moreover, T10 up-regulated mGlu5 expression decreased by LPS through enhancing mRNA expression and protein stability. T10 also reversed the reduction in mGlu5 membrane localization and modulated receptor-mediated mitogen-activated protein kinase activity induced by LPS. Pharmacological inhibition of signaling molecules increased nitric oxide level and inducible nitric oxide synthase (iNOS), tumor necrosis factor-α, and interleukin (IL)-1β and -6 transcript levels that were downregulated by treatment with T10. Consistent with these in vitro findings, blocking mGlu5 attenuated the anti-inflammatory effects of T10 in an LPS-induced PD model and blocked the decreases in the number and morphology of ionized calcium binding adaptor molecule 1-positive microglia and LPS-induced iNOS protein expression caused by T10 treatment. Besides, mGlu5 mediated the effect of T10 on microglia-induced astrocyte activation in vitro and in vivo. The findings provide evidence for a novel mechanism by which mGlu5 regulates T10-inhibited microglia activation via modulating protein expression of the receptor and its intracellular signaling. The study might contribute to the biological effects of Chinese herbs as an approach for protecting against neurotoxicity in PD.

Six undescribed pentacyclic triterpenoids including four triterpenoid aglycones, 1β,2a,3β,23-tetrahydroxyurs-12-en-28-ursolic acid, 2a,3a,6β,19α,23-pentahydroxyurs-12-en-28-ursolic acid, 2α,3α,20β,23-tetrahydroxyurs-12-en-28-ursolic acid and 1β,2a,3β,23-tetrahydroxyurs-12,20(30)-dien-28-ursolic acid, and two triterpenoid glucosides, 2a,3a,23-trihydroxy-12,20(30)-dien-28-ursolic acid 28-O-β-d-glucopyranoside and 1-oxo-3β,23-dihydroxyolean-12-en-28-oic acid 28-O-β-d-xylopyranoside, along with 5 known triterpenoids were isolated from a CH3Cl-soluble extract of the leaves of Cyclocarya paliurus. Their structures were established on the basis of chemical and spectroscopic approaches. These compounds were assessed for their antioxidant effects on FFA-induced hepatic steatosis in HepG2 cells. The results revealed that three saponins and two aglycones markedly increased SOD activity and reduced MDA level.

Four new C-11 monosaccharide attached dammarane triterpenoid glycosides cypaliurusides S V (1–4), along with nine known dammarane triterpenoid glycosides (5–13) were isolated from a CHCl3-soluble extract of the leaves of Cyclocarya paliurus. All characterized compounds were assayed for their cytotoxicities against HepG2 cells and 10 compounds were evaluated for the agonistic effects on sirtuin1 (SIRT1). The results showed that compounds 1, 5 and 6 were strongly cytotoxic in HepG2 cell line. Two dammarane triterpenoid glycosides 3 and 10 exhibited agonistic activities on SIRT1 with IC50 of 10 μM and 20 μM, respectively.

The capture and reuse of NH 4 + is an ideal solution to treat NH 4 + -containing wastewater. The capture and reuse process needs to be clean and cost-effective. Currently, however, there are many obstacles, particularly in the availability, cost, and recovery of the chemical sources required. Here, we demonstrate a clean and efficient method to capture and recycle NH 4 + by a dolomite-aided struvite precipitation process. Dolomite calcined carefully in CO 2 atmosphere was used as a Mg source to react with PO 4 3− (KH 2 PO 4 ) and NH 4 + in model wastewater (2000 mg L −1 NH 4 + ). The precipitation was performed at nMg 2+ :nNH 4 + :nPO 4 3− = 1:1:1.2 and pH = 8.0 for 2 h; 89.7% of NH 4 + was recovered in the form of struvite precipitate. The competition between K + and NH 4 + in the model wastewater led to the formation of K-struvite (MgKPO 4 ·6H 2 O) and struvite (MgNH 4 PO 4 ·6H 2 O). The formation of K-struvite resulted in a decrease in the NH 4 + removal rate. When struvite was heated at 110 °C for 4 h, the NH 4 + release rate from the thermolysis reached 75.7%. Thermolysis readily occurred as an unstable Ca 2+ -CO 3 2- -NH 4 + system formed in the mixture of MgNH 4 PO 4 ·6H 2 O and CaCO 3 . The elements Mg and P that were obtained during the struvite precipitation–thermolysis–reprecipitation process can be repeatedly used. After 6 cycles, under the conditions pH = 9.0, nMg 2+ :nNH 4 + :nPO 4 3− = 1:1:1 and reaction time of 2 h, up to 78.3% of NH 4 + was removed from the model wastewater.

Numerous epidemiological studies suggested that there is a variable cancer risk in patients with Parkinson's disease (PD). However, the underlying mechanisms remain unclear. In the present study, the role of metabotropic glutamate receptor 5 (mGluR5) has been investigated in 6-hydroxydopamine (6-OHDA)-induced PD combined with liver cancer both in vitro and in vivo. We found that PD cellular model from 6-OHDA-lesioned MN9D cells suppressed the growth, migration, and invasion of Hepa1-6 cells via down-regulation of mGluR5-mediated ERK and Akt pathway. The application of 2-methyl-6-(phenylethyl)-pyridine and knockdown of mGluR5 further decreased the effect on Hepa-1-6 cells when co-cultured with conditioned media. The effect was increased by (S)-3,5-dihydroxyphenylglycine and overexpression of mGluR5. Moreover, more release of glutamate from 6-OHDA-lesioned MN9D cells suppressed mGluR5-mediated effect of Hepa1-6 cells. Application of riluzole eliminated the increased glutamate release induced by 6-OHDA in MN9D cells and aggravated the suppressive effect on Hepa-1-6 cells. In addition, the growth of implanted liver cancer was inhibited in 6-OHDA induced PD-like rats, and was associated with increased glutamate release in the serum and down-regulation of mGluR5 in tumor tissue. Collectively, these results indicate that selective antagonism of glutamate and mGluR5 has a potentially beneficial effect in both liver cancer and PD, and thus may provide more understanding for the clinical investigation and further an additional therapeutic target for these two diseases.

In the search for novel solid acid catalysts for the gas-phase catalytic dehydration of glycerol to acrolein, sepiolite clay minerals were activated with hydrochloric acid to produce acid-activated sepiolite catalysts. The effects of the activation process on the texture and acidity of acid-activated sepiolite, and their roles in gas-phase catalytic dehydration of glycerol to acrolein, were then investigated. A series of acid-activated sepiolite catalysts were made from purified natural sepiolite treated with hydrochloric acid of different concentrations (0–8 mol/L) at 80 °C, followed by drying at 100 °C. The acid-activated sepiolite catalyst samples were characterized using X-ray diffraction, X-ray fluorescence analysis, scanning electronic microscopy, thermogravimetric analysis, N2 adsorption-desorption isotherms, NH3-temperature-programmed desorption. Fourier transform-infrared spectroscopy and pyridine adsorption followed by in situ infrared spectroscopy. The catalytic performances in gas-phase catalytic dehydration of glycerol to acrolein were studied using a vertical fixed-bed reactor. Typically, a glycerol conversion of 92.9%, an acrolein selectivity of 59.4%, and a yield of acrolein of 55.2% were achieved when the gas-phase catalytic dehydration of glycerol was conducted over HCl (2 mol/L) acid-activated sepiolite catalyst at 320 °C with an aqueous glycerol solution (20 wt.%) at a rate of 0.10 ml/min as the feedstock and air as the carrier gas at a flow rate of 20 ml/min. The activation of the sepiolite with hydrochloric acid followed by drying can eliminate a portion of the zeolitic water in the tunnels, remove part of the magnesium and aluminum cations in the octahedral sheets of sepiolite, unbundle aggregated sepiolite fibers, and partly break the Si-O-Si in the tetrahedral sheets of sepiolite. These factors increased the porosity, the specific surface area and the acidity of the sepiolite. The partial leaching of magnesium and aluminum ions in the octahedral sheet generated octahedral vacancies and created Mg O Al+ sites in the octahedral sheets, thereby increasing the amount of Lewis acid sites. Along with penetration of H+ cations into the interlayered space of the sepiolite by an ion-exchange reaction and physicochemical surface adsorption, some breakage of the Si O Si bonds in the tetrahedral sheets generated more Si O H+, thus increasing the quantity of Bronsted acid sites. Furthermore, the strength of acidity of the hydrochloric acid-activated sepiolite was significantly changed after the acid activation. The medium-strong Bronsted acid sites (Si O H+ and H+ adsorbed on the surface) appeared to be beneficial for the high conversion of glycerol and the selectivity to acrolein. Lewis acid sites could facilitate the formation of acetol. Under the reaction conditions mentioned above, the coking on the hydrochloric acid-activated sepiolite catalysts readily occurred, rapidly leading to the decrease of the yield of acrolein.

The objective of this study was to evaluate the effects on chemical and microbiological properties of paddy soil of short-term biochar, straw, and chemical fertilizers compared with chemical fertilization alone. Five soil fertilization treatments were evaluated: regular chemical fertilizers (RF), straw+regular chemical fertilizers (SRF), straw biochar+regular chemical fertilizers (SCRF), bamboo biochar (BC)+regular chemical fertilizers (BCRF), and straw biochar+70% regular chemical fertilizers (SC+70%RF). Their effects were investigated after approximately 1.5 years. The soil pH and cation exchange capacity (CEC) were significantly higher in biochar-treated soils. The soil phosphorous (P) and potassium (K) contents increased with biochar application. The soil Colwell P content was significantly increased with the addition of straw biochar in the treatments of SCRF and SC+70%RF. The oxygen (0):carbon (C) ratio doubled in BC picked from the soil. This indicated that BC underwent a significant oxidation process in the soil. The denaturing gradient gel electrophoresis (DGGE) fingerprints of microbial communities differed among the treatments. Soils with added biochar had higher Shannon diversity and species richness indices than soils without biochars. The results suggest that biochar can improve soil fertility.

Anorectal melanoma is a rare disease that carries a poor prognosis. To date, limited genetic analyses confirmed KIT mutations as a recurrent genetic event similar to other mucosal melanomas, occurring in up to 30% of anorectal melanomas. Importantly, a subset of tumors harboring activating KIT mutations have been found to respond to c-Kit inhibitor-based therapy, with improved patient survival at advanced tumor stages. We performed comprehensive targeted exon sequencing analysis of 467 cancer-related genes in a larger series of 15 anorectal melanomas, focusing on potentially actionable variants based on gain- and loss-of-function mutations. We report the identification of oncogenic driver events in the majority (93%) of anorectal melanomas. These included variants in canonical MAPK pathway effectors rarely observed in cutaneous melanomas (including an HRAS mutation, as well as a BRAF mutation resulting in duplication of threonine 599), and recurrent mutations in the tumor suppressor NF1 in 20% of cases, which represented the second-most frequently mutated gene after KIT in our series. Furthermore, we identify SF3B1 mutations as a recurrent genetic event in mucosal melanomas. Our findings provide an insight into the genetic diversity of anorectal melanomas, and suggest significant potential for alternative targeted therapeutics in addition to c-Kit inhibitors for this melanoma subtype.

Neuroendocrine carcinomas (NECs) of the colorectum are rare but highly aggressive neoplasms. These tumors show some shared genetic alterations with colorectal adenocarcinomas, and most of them have adjacent glandular adenoma or adenocarcinoma components. However, genetic data on colorectal NECs still are sparse and insufficient for definite conclusions regarding their molecular origin. Based on morphological characterization, panel and whole-exome sequencing, we here present results from an in-depth analysis of a collection of 15 colorectal NECs with glandular components, 10 of which by definition were mixed adenoneuroendocrine carcinomas (MANECs). Among shared genetic alterations of both tumor components, we most frequently found TP53, KRAS and APC mutations that also had highest allele frequencies. Mutations exclusive to glandular or neuroendocrine components outnumbered shared mutations but occurred at lower allele frequencies. Our findings not only provide additional evidence for a common clonal origin of colorectal NECs and adjacent glandular tumor components, but strongly suggest their development through the classical adenoma-carcinoma sequence. Moreover, our data imply early separation of glandular and neuroendocrine components during malignant transformation with subsequent independent mutational evolution.

Although there are numerous strategies to counteract the death of dopaminergic neurons in Parkinson's disease (PD), there are currently no treatments that delay or prevent the disease course, indicating that early protective treatments are needed. Targeting axonal degeneration, a key initiating event in PD, is required to develop novel therapies; however, its underlying molecular mechanisms are not fully understood. Here, we studied axonal degeneration induced by 6-hydroxydopamine (6-OHDA) in vitro and in vivo. We found that metabotropic glutamate receptor 5 (mGluR5) expression increased during 6-OHDA-induced axonal degeneration in primary neurons and that blockade of mGluR5 by its antagonists 2-methyl-6-(phenylethynyl)-pyridine (MPEP) and 3-[(2-methyl-1, 3-thiazol-4-yl) ethynyl]-pyridine (MTEP) almost completely attenuated the degenerative process in vitro. Furthermore, a rapid increase in intra-axonal calcium levels following 6-OHDA treatment was visualized using a calcium-sensitive fluorescence probe and a calcium chelator prevented the axonal degenerative process induced by 6-OHDA in vitro, whereas application of the mGluR5 antagonist MPEP partially attenuated the increase in intra-axonal calcium. The screening of calcium targets revealed that calpain activation and an increase in phosphorylated extracellular signal-regulated kinase (p-ERK) were calcium dependent during 6-OHDA-induced axonal degeneration in vitro. Consistent with these in vitro findings, blockade of mGluR5 with MPEP attenuated the degeneration of dopaminergic axons induced by 6-OHDA injection into the striatum prior to soma death in the early stage of PD in an in vivo animal model. In addition, MPEP inhibited the increase in mGluR5 expression levels, calpain activation and the elevation of p-ERK in the striatum triggered by 6-OHDA injection in vivo. Taken together, these data identify an mGluR5-calcium-dependent cascade that causes axonal degeneration, and suggest that mGluR5 antagonists could provide effective therapy to prevent the disease process of PD.

Fly ash is the fine solid particulate residue driven out of the boiler with the flue gases in coal-fired power plants. Now it can be used for making geopolymer which acts as a cement-like product. The geopolymer technology provides an alternative good solution to the utilization of fly ash with little negative impact on environment. This review summarizes and examines the scientific advances in the preparation, properties and applications of fly ash-based geopolymer. The production of fly ash-based geopolymer is mainly based on alkali activated geopolymerization which can occur under mild conditions and is considered as a cleaner process due to much lower CO 2 emission than that from the production of cement. The geopolymerization can trap and fix the trace toxic metal elements from fly ash or external sources. The Si/Al ratios, the type and the amount of the alkali solution, the temperature, the curing conditions, and the additives are critical factors in a geopolymerization process. The mechanical performances of the fly ash-based geopolymer, including compressive strength, flexural and splitting tensile strength, and durability such as the resistance to chloride, sulfate, acid, thermal, freeze-thaw and efflorescence, are the primary concerns. These properties of fly ash-based geopolymer are inherently dependent upon the chemical composition and chemical bonding and the porosity. The mechanical properties and durability can be improved by fine tuning Si/Al ratios, alkali solutions, curing conditions, and adding slag, fiber, rice husk-bark ash and red mud. Fly ash-based geopolymer is expected to be used as a kind of novel green cement. Fly ash-based geopolymer can be used as a class of materials to adsorb and immobilize toxic or radioactive metals. The factors affecting the performances of fly ash-based geopolymer concrete, in particular aggregate, are discussed. For future studies on fly ash-based geopolymer, further enhancing mechanical performance, scaling up production and exploring new applications are suggested.

Montmorillonite-hydrochar nanocomposite is of great significance in biofuel production, energy storage and conversion, catalysis, environmental protection, and biology and medicine. To understand how montmorillonite facilitates the conversion of cellulose into hydrochar, it is essential to accurately control the properties and structure of hydrochar and its nanocomposites for technological applications. In this work, hydrochar and montmorillonite-hydrochar nanocomposites were produced by hydrothermal carbonization of cellulose at 200 °C for 2–24 h in the absence and the presence of montmorillonite. The roles of montmorillonite in the formation of hydrochar were explored in detail. Results showed that montmorillonite acted as a catalyst, an adsorbent, and an inorganic template for hydrochar formation. The formation process of montmorillonite-hydrochar nanocomposite can be divided into two stages: first, the hydrolysis of cellulose yielding degraded liquid products (e.g. glucose, fructose, and organic acids); second, the formed liquid products were adsorbed by montmorillonite, which catalyzed condensation and aromatization to form hydrochar particles on the surface of montmorillonite.

Mucinous cystic neoplasm (MCN) is characterized by mucinous epithelial lining and subepithelial ovarian-like stroma and is a precursor to invasive carcinoma. Type 1 autoimmune pancreatitis (AIP), on the other hand, is a form of IgG4-related disease (IgG4-RD) and can present as a pancreatic mass. There is no known causal relationship between these two entities in the current literature. Here, we report a case of a middle-aged female patient who was found to have an enlarging pancreatic mass and subsequently underwent a distal pancreatectomy. Gross examination of the pancreatectomy specimen showed a 4.3 cm mass with a central 3.4 cm cystic component, confirmed to be an MCN on microscopy. The immediately adjacent peritumoral tissue showed storiform fibrosis, obliterative phlebitis, and focal IgG4-positive plasma cell–rich infiltrates (≥75/HPF) with an IgG4+/IgG+ ratio of 80%. The background pancreas was histologically normal and these characteristic AIP findings were limited to the pericystic tissue. Clinical workup with CT scan and MRI did not show any evidence of systemic IgG4-RD, nor was the serum IgG4 level ever elevated. To our knowledge, this is only the second reported case of a pancreatic MCN associated with peritumoral AIP-like changes in a patient without systemic IgG4-RD. Awareness of these potential AIP-like changes surrounding pancreatic neoplasms may be helpful in preventing misdiagnosis in needle biopsy and aspiration specimens. Keywords: Type 1 autoimmune pancreatitis, IgG4-related disease, IgG4-RD, IgG4-related pancreatitis, Mucinous cystic neoplasm, MCN, Sclerosing pancreatitis

For the development of efficient solid acid catalysts for the catalytic dehydration of glycerol to acrolein, catalysts made from montmorillonitic clay activated by sulfuric acid were investigated. Montmorillonite was activated in diluted sulfuric acid in the concentration range of 5–40 wt.%. The effects of sulfuric acid treatment on the structure of the montmorillonite were characterized by X-ray diffraction, measurements of acidity, N2 adsorption–desorption isotherms, and Fourier transform infrared spectroscopy. The catalytic behavior of sulfuric acid-activated montmorillonite catalysts in the gas-phase dehydration of glycerol were investigated under varying conditions, including the reaction temperature, the feed rate, and the concentration of glycerol. After montmorillonitic clay was activated by sulfuric acid, the layered structural features of montmorillonite remained nearly intact. Ca2 +-montmorillonite was changed to H+-montmorillonite by ion exchange reaction during activation. The optimal catalytic glycerol dehydration reaction conditions were found to be: temperature at 320 °C, liquid hourly space velocity (LHSV) = 18.5 h− 1, concentration of glycerol solution = 10 wt.%, and the flow rate of N2 carrier gas = 10 mL/min. A conversion of 54.2% of glycerol and a yield of 44.9 wt.% acrolein were achieved over the montmorillonite catalyst activated by an aqueous 10 wt.% sulfuric acid solution. The H+ in the interlayer space of acid-activated montmorillonite catalysts played a critical role in the catalytic dehydration of glycerol. The temperature, the LHSV, and the concentration of glycerol affected the performance of the catalysts through their influence on the reaction mechanism, the contact time, and the reaction equilibrium.

Transcriptional changes accompany induced resistance of plants against insect feeding, and monitoring transcriptional reorganization triggered in response to herbivory is an essential step for deciphering the molecular basis of the resistance. To isolate up-regulated genes in tea leaves under mild infestation of green leafhopper (Empoasca vitis Gothe), a subtractive cDNA library was constructed using the suppression subtractive hybridization strategy. Subsequent differential screening, DNA sequencing analyses, and real-time PCR analyses identified 21 unique genes up-regulated by leafhopper feeding from the library. Among the genes likely to be directly involved in defense, those encoding pathogen-related protein and type 2 ribosome-inactivating protein were found in the subtractive library. In addition, genes involved in the CBL (calcineurin B-like)–CIPK (CBL-interacting protein kinases) pathway, cell wall fortification, secondary metabolite synthesis, water transport, and oxidative stress protection were identified in our experiment. Besides, a number of genes that have not been previously associated with defense responses against insects were isolated.

Study objectives To examine the lung defensemechanisms of both young and aged rats before and after pulmonarychallenge with a bacterial pathogen. Design Male Fischer 344 rats, either 2.5 months or 20 months of age, wereintratracheally inoculated with 5 × 10 3 ,5 × 10 4 , or 5 × 10 5 Listeriamonocytogenes , and the effects on mortality, lung inflammation, pulmonary bacterial clearance, alveolar macrophage (AM) function, and T-lymphocyte characterization were determined. Measurementsand results In noninfected control animals, the older rats hadlower numbers of AMs on lavage and a lower percentage of total T, CD4+, and CD8+ cells. No difference was observed between noninfected youngand old rats in AM function, assessing both chemiluminescence andnitric oxide (NO) production. After bacterial challenge, aged ratsexhibited an increase in mortality, pulmonary infection, and edema, andlung lesions, which were more extensive than those observed in theyounger rats. Interestingly, AM chemiluminescence was enhanced, while AM NO, a highly important antibacterial defense product, was abrogatedin the aged rats as compared to the young rats. Conclusions This study demonstrated that advanced age isassociated with alterations in lung defense mechanisms and increasedsusceptibility to pulmonary bacterial infection marked by elevatedmortality, slowed pulmonary bacterial clearance, and altered AMfunction, specifically a decrease in NO production. These observationsare indicative of reduced pulmonary defense function in an olderpopulation of rats.

Dichloroacetic acid (DCA) and chloroform (CHCl 3 ) are both major by-products of drinking water chlorination and DCA increases the hepatotoxicity of CHCl 3 . In this study, we further characterized this effect and investigated DCA-induced alterations of CHCl 3 disposition and metabolism as a possible mechanism for this interaction. Both adult male and female Sprague-Dawley rats were gavaged with three doses (09:00, 16:00 and 09:00 the next morning) of DCA (each 2.45 mmol/kg), then challenged with an i.p. injection of CHCl 3 (3.12 or 9.35 mmol/kg). Hepatic damage was assessed 24 h after CHCl 3 administration as increased alanine aminotransferase (ALT), ornithine carbomyl transferase (OCT) and bilirubin in plasma. In a separate experiment, rats were pretreated with DCA or were given 14 CHCl 3 at the same dosages. The disposition of 14 C in various tissues and covalent binding of 14 CHCl 3 -derivatives to liver proteins and lipids were determined 1 h later. CHCl 3 -induced hepatotoxicity was significantly more severe in DCA-pretreated groups. ALT and OCT were more markedly elevated in DCA+CHCl 3 (3.12 mmol/kg) groups than NaCl+CHCl 3 animals. Plasma bilirubin content was elevated only in DCA+CHCl 3 groups and females were more susceptible to this effect. The responses of rats to DCA treatment were somewhat gender-different. DCA treatment increased total cytochrome P450 in females, but not in males. Hepatic glutathione concentration was elevated in males after DCA treatment, but not in females. In the present study we confirmed that DCA pretreatment potentiates the CHCl 3 -hepatotoxicity of both male and female rats. However, there was little evidence that DCA pretreatment significantly affected CHCl 3 disposition or increased CHCl 3 binding in vivo.

Previously, we showed that the pretreatment of dichloroacetic acid (DCA) increased CHCl3-induced hepatotoxicity in vivo and CHCl3 metabolism in vitro in both male and female rats. The effects of DCA on hepatic cytochrome P450-dependent monooxygenases were studied in this experiment. Male and female Sprague-Dawley rats were treated with DCA (each 2.45 mmol/kg) three times (9 AM, 4 PM, and 9 AM) and hepatic microsomes were prepared 3 hr after the last treatment (the same procedure as previous studies). After DCA treatment, the total content of cytochrome P450 (0.67 nmol/mg protein vs 0.79) and the activity of NADPH-dependent cytochrome P450 reductase (227 nmol/mg protein/min vs 332) were significantly increased in female rats, but they were unchanged in males (0.99 vs 0.98 for P450; 315 vs 311 for reductase). Induction of CYP2E1 was observed in both sexes, evidenced as increased activities of aniline and p-nitrophenol hydroxylases and increased CYP2E1 protein amount determined by immunoblot assay. In contrast, the CYP2B-related activity (dealkylation of pentoxyresorufin) and immunoreactive protein did not increase after DCA treatment in either males or females. The activity of ethoxyresorufin dealkylase was decreased in DCA-treated males compared to their controls (310 pmol/mg protein/min vs 229, p0.05), but it was not significantly affected in females. These data demonstrate that DCA treatment of both male and female rats altered the population of hepatic cytochrome P450. The results support the hypothesis that DCA increases CHCl3 metabolism, and therefore hepatotoxicity, by inducing CYP2E1.

Physiologically based pharmacokinetic (PBPK) models developed from gas uptake experiments have been used to estimate metabolic parameters for volatile organic compounds. Due to the potential application of PBPK models to estimate metabolic bioactivation constants in humans, it is important to understand the complex nature of these models and the resulting estimates. Adult male F344 rats (165-205 g) were individually exposed to carbon tetrachloride (CC14) in gas uptake systems. Three rats at each concentration were exposed for 6 hr to initial concentrations of 25, 100, 250, and 1000 ppm CCl4. Partition coefficient determinations were performed by the vial equilibration technique and used as model inputs. Computer optimizations with the means of each initial chamber concentration at each time point resulted in an estimate of Vmax of 0.11 mg/hr (Vmaxc = 0.37 mg/hr/kg) and Km of 1.3 mg/liter. To determine the effect of individual animal variation in Vmax optimizations were also performed with the mean ± SD, resulting in Vmax estimates of 0.09 and 0.12 mg/hr, respectively. Similar analysis resulted in Km estimates of 0.98 and 1.58 mg/liter. The results of the sensitivity analysis were concentration dependent for CCl4. These results show Vmax and Km to be most accurately detected at lower initial chamber concentrations. Results of the sensitivity analysis at the lowest concentration established the following model input hierarchy: blood to air partition > fat partition and fat volume fraction > slowly perfused partition, ventilation rate, cardiac output, fat blood flow percentage > liver blood flow percentage and slowly perfused blood flow percentage. Further sensitivity analysis determined Vmax and Km to be highly correlated when using gas uptake technology and point to the need to an independent estimate for either constant. In summary, the application of sensitivity analysis to PBPK modeling resulted in an increased understanding of factors governing the estimation of metabolic parameters.