Hepatic Biotransformation of Renal Clearable Gold Nanoparticles for Noninvasive Detection of Liver Glutathione Level via Urinalysis.

Renal clearable nanoparticles have been drawing much attention as they can avoid prolonged accumulation in the body by efficiently clearing through the kidneys. While much effort has been made to understand their interactions within the kidneys, it remains unclear whether their transport could be influenced by other organs, such as the liver, which plays a crucial role in metabolizing and eliminating both endogenous and exogenous substances through various biotransformation processes. Here, by utilizing renal clearable IRDye800CW conjugated gold nanocluster (800CW4-GS18-Au25) as a model, we found that although 800CW4-GS18-Au25 strongly resisted serum-protein binding and exhibited minimal accumulation in the liver, its surface was still gradually modified by hepatic glutathione-mediated biotransformation when passing through the liver, resulting in the dissociation of IRDye800CW from Au25 and biotransformation-generated fingerprint message of 800CW4-GS18-Au25 in urine, which allowed us to facilely quantify its urinary biotransformation index (UBI) via urine chromatography analysis. Moreover, we observed the linear correlation between UBI and hepatic glutathione concentration, offering us a noninvasive method for quantitative detection of liver glutathione level through a simple urine test. Our discoveries would broaden the fundamental understanding of in vivo transport of nanoparticles and advance the development of urinary probes for noninvasive biodetection.

Photo-on-Demand In Situ One-Pot Synthesis of Carbonate Esters from Tetrachloroethylene.

The present study reports a novel one-pot synthesis of carbonate esters with photo-oxidized tetrachloroethylene (TCE). Acyclic and cyclic alkyl carbonate esters could be synthesized through base-promoted condensation reactions of alcohols with the photo-oxidized TCE that was prepared by irradiation with UV-C or visible light under O2 or O2/Cl2 (∼4%) bubbling, respectively. Cyclic carbonate esters could also be synthesized from a solution of TCE and the ethylene glycol derivative by irradiation of UV-C light under O2 bubbling. With respect to the reaction mechanism, the photochemical oxidation of TCE mainly provides the highly toxic and corrosive trichloroacetyl chloride (TCAC), which then reacts in situ with the alcohol to give the corresponding trichloroacetic acid ester (TCAE). The subsequent intermolecular or intramolecular base-catalyzed condensation reaction of TCAE with or without the addition of alcohol leads to elimination of CHCl3 in the corresponding carbonate ester. The present reactions enable the in situ one-pot synthesis of a variety of alkyl carbonate esters under mild conditions without the direct handling of TCAC. This is beneficial in terms of safety, cost, and reduced environmental impact.

Separation and quantification of Azvudine in plasma of patients with COVID-19 using LC-MS/MS.

Azvudine (FNC) is a new drug conditionally approved in 2022 for the treatment of coronavirus disease 2019 (COVID-19) in China. However, the exposure level of FNC in COVID-19 patients in clinical practice is still obscure, and there is no liquid chromatography-tandem mass spectrometry (LC-MS/MS) or LC method reported for quantifying the FNC. In this study, a simple, fast, and reliable LC-MS/MS method using L-phenylalanine-D5 (Phe-D5) as the internal standard (IS) was developed for the quantification of FNC in plasma from COVID-19 patients. After simple protein precipitation with methanol, the analyte in the supernatant was separated on Waters Atlantis® T3 (2.1 ×100 mm, 3.0 µm) column with the mobile phase consisting of acetonitrile (ACN) - aqueous solution (containing 0.03% heptafluorobutyric acid and 0.2% formic acid). The mobile phase was delivered at 0.3 mL/min in an isocratic elution program (15:85, V: V). The linear relationship of FNC was good within the calibration range of 2.0 - 2000.0 ng/mL, with the recovery of FNC ranging from 81.37% to 103.31% and the matrix effect was 94.77%- 109.83%. The short-term, long-term, and freeze-thaw stability of the FNC assessed in method was acceptable, and all other items met the requirements of validation of the biological analytical method. Finally, the method was applied to detect the exposure level of FNC in plasma samples from patients diagnosed with COVID-19, and the results, which are within the linear range of the method, showed huge inter-individual variation, supporting the significance of therapeutic drug monitoring of FNC.

Photo-on-Demand In Situ Phosgenation Reactions That Cross Three Phases of a Heterogeneous Solution of Chloroform and Aqueous NaOH.

Here, we report a novel photo-on-demand in situ phosgenation reaction that crosses three phases of a heterogeneous solution of chloroform (CHCl3) and aqueous NaOH containing an aryl alcohol or amine. This reaction system enables the safe, convenient, and inexpensive synthesis of carbonate esters, polycarbonates, and N-substituted ureas from aryl alcohols, aryl diols, and primary/secondary amines, respectively, on a practical scale and with good yield. The photochemical oxidation of CHCl3 to phosgene (COCl2) occurs upon irradiation with UV light from a low-pressure mercury lamp of both the gas and liquid phases of the reaction system under O2 bubbling of the vigorously stirred sample solution. The following reaction mechanisms are suggested: The aryl alcohol reacts in situ with the generated COCl2 at the interfaces of the organic/aqueous phases and aqueous/gas phases, in competition with the decomposition of COCl2 due to hydrolysis. Nucleophilicity and hydrophilicity are enhanced by the formation of aryl alkoxide ion through the reaction with NaOH, whereas the reaction of amine proceeds through neutralization of the generated HCl by the aqueous NaOH.

Dual-energy computed tomography in reducing the effective radiation dose of computed tomography urography in patients with urinary calculi.

Background: To evaluate the diagnostic performance of split-bolus single-phase dual-energy computed tomography (DECT) with virtual non-contrast computed tomography (VNCT) compared to three-phase computed tomography (CT) urography in patients with urinary calculi, and to examine the performance of split-bolus single-phase DECT when reducing the effective dose. Methods: A total of 48 patients with abdominal pain or hematuria suggestive of unilateral urinary calculi were enrolled and randomly divided into the experimental and control groups, with 24 cases in each group. Patients in the experimental group underwent split-bolus single-phase DECT to obtain a mixed nephrographic excretory phase. Patients in the control group accepted a single-bolus three-phase CT urography scan (non-contrast, nephrographic phase, and excretory phase). The CT values and the contrast-to-noise ratio (CNR) of 7 segments of the urinary tract were measured and compared between the two groups by using the Mann-Whitney U test. The dose-length product (DLP) and effective dose of each patient were compared between the two groups using an independent t-test. Results: Among all 48 patients, 35 calculi were detected in the experimental group (n=24), and 47 calculi were detected in the control group (n=24). There was no significant difference between the two groups in both CT value measurements and the CNR. The mean DLP and mean effective dose of the experimental group were significantly lower than those of the control group, and the effective dose in the experimental group was decreased by 40% compared with the control group. Conclusions: The application of DECT combined with split-bolus nephrographic excretory phase CT urography can reveal the urinary calculi covered by a contrast medium and also reduce the effective dose exposure to patients.

Photo-on-Demand In Situ Synthesis of N-Substituted Trichloroacetamides with Tetrachloroethylene and Their Conversions to Ureas, Carbamates, and Polyurethanes.

N-substituted trichloroacetamides (NTCAs), which serve as blocked isocyanates, were synthesized in ∼97% yields by in situ photo-on-demand trichloroacetylation of amines with tetrachloroethylene (TCE). The reactions were performed by photo-irradiation of TCE solutions containing an amine under O2 bubbling over 70 °C with a low-pressure mercury lamp. TCE underwent photochemical oxidation to afford trichloroacetyl chloride having high toxicity and corrosivity, which then reacts in situ with the amine to afford NTCA. Compared with conventional NTCA synthesis with hexachloroacetone, the present reaction has the advantage of being widely applicable to a variety of amines, even those with low nucleophilicity such as amides, fluorinated amines, and amine HCl salts. NTCAs could be converted to the corresponding N-substituted ureas and carbamates through base-catalyzed condensation with amines and alcohols, respectively, with the elimination of CHCl3. The reaction may proceed by the initial formation of isocyanate and its subsequent addition reaction with the amine or alcohol. This photochemical reaction also enables the synthesis of fluorinated NTCAs, which accelerate the reactions, and realizes the synthesis of novel fluorinated chemicals including polyurethanes.

Photo-On-Demand Synthesis of α-Amino Acid N-Carboxyanhydrides with Chloroform.

Amino acid N-carboxyanhydrides (NCAs) are conventionally synthesized from α-amino acids and phosgene. The present study reports in situ photo-on-demand phosgenation reactions of amino acids with CHCl3 for synthesizing NCAs. A series of NCAs were obtained on a gram scale upon photo-irradiation of a mixture solution of CHCl3 and CH3CN containing an amino acid at 60-70 °C under O2 bubbling. This method presents a safe and convenient reaction controlled by light without special apparatuses and reagents.

Identification of Abietane-Type Diterpenoids and Phenolic Acids Biosynthesis Genes in Salvia apiana Jepson Through Full-Length Transcriptomic and Metabolomic Profiling.

Salvia apiana (S. apiana) Jepson is a medicinal plant that is frequently used by the Chumash Indians in southern California as a diaphoretic, calmative, diuretic, or antimicrobial agent. Abietane-type diterpenoids (ATDs) and phenolic acids (PAs) are the main bioactive ingredients in S. apiana. However, few studies have looked into the biosynthesis of ATDs and PAs in S. apiana. In this study, using metabolic profiling focused on the ATDs and PAs in the roots and leaves of S. apiana, we found a distinctive metabolic feature with all-around accumulation of ATDs, but absence of salvianolic acid B. To identify the candidate genes involved in these biosynthesis pathways, full-length transcriptome was performed by PacBio single-molecule real-time (SMRT) sequencing. A total of 50 and 40 unigenes were predicted to be involved in ATDs and PAs biosynthesis, respectively. Further transcriptional profile using Illumina HiSeq sequencing showed that the transcriptional variations of these pathways were consistent with the accumulation patterns of corresponding metabolites. A plant kingdom-wide phylogenetic analysis of cytochromes (CYPs) identified two CYP76AK and two CYP76AH subfamily genes that might contribute for the specific ATDs biosynthesis in S. apiana. We also noticed that the clade VII laccase gene family was significantly expanded in Salvia miltiorrhiza compared with that of S. apiana, indicating their involvements in the formation of salvianolic acid B. In conclusion, our results will enable the further understanding of ATDs and PAs biosynthesis in S. apiana and Salvia genus.

Direct Syntheses of Diphenylmethanol Derivatives from Substituted Benzenes and CHCl3 through Friedel-Crafts Alkylation and Post-Synthetic Hydrolysis or Alcoholysis Catalyzed by Alumina.

The present study reports an innovative finding that alumina containing water or primary alcohol catalyzes the hydrolysis or alcoholysis, respectively, of the product formed through AlCl3 -mediated Friedel-Crafts alkylation of methyl-substituted benzenes and CHCl3 . The former and later reactions mainly provided hydroxy- and alkoxy-substituted diarylmethanes, respectively, while the reference reactions without alumina provided bisarylchloromethane. This method enables the selective syntheses of diphenylmethanol derivatives with very simple procedures, without expensive reagents and apparatuses. Furthermore, the alumina used in the reaction could be recycled by washing with water and subsequent drying. From the viewpoint of material recycling, this function is very important for the development of sustainable chemical reactions.

Molecular Mapping and Analysis of an Excellent Quantitative Trait Loci Conferring Adult-Plant Resistance to Stripe Rust in Chinese Wheat Landrace Gaoxianguangtoumai.

The Chinese wheat landrace "Gaoxianguangtoumai" (GX) has exhibited a high level of adult-plant resistance (APR) to stripe rust in the field for more than a decade. To reveal the genetic background for APR to stripe rust in GX, a set of 249 F6:8 (F6, F7, and F8) recombinant inbred lines (RILs) was developed from a cross between GX and the susceptible cultivar "Taichung 29." The parents and RILs were evaluated for disease severity at the adult-plant stage in the field by artificial inoculation with the currently predominant Chinese Puccinia striiformis f. sp. tritici races during three cropping seasons and genotyped using the Wheat 55K single-nucleotide polymorphism (SNP) array to construct a genetic map with 1,871 SNP markers finally. Two stable APR quantitative trait loci (QTL), QYr.GX-2AS and QYr.GX-7DS in GX, were detected on chromosomes 2AS and 7DS, which explained 15.5-27.0% and 11.5-13.5% of the total phenotypic variation, respectively. Compared with published Yr genes and QTL, QYr.GX-7DS and Yr18 may be the same, whereas QYr.GX-2AS is likely to be novel. Haplotype analysis revealed that QYr.GX-2AS is likely to be rare which presents in 5.3% of the 325 surveyed Chinese wheat landraces. By analyzing a heterogeneous inbred family (HIF) population from a residual heterozygous plant in an F8 generation of RIL, QYr.GX-2AS was further flanked by KP2A_36.85 and KP2A_38.22 with a physical distance of about 1.37Mb and co-segregated with the KP2A_37.09. Furthermore, three tightly linked Kompetitive allele-specific PCR (KASP) markers were highly polymorphic among 109 Chinese wheat cultivars. The results of this study can be used in wheat breeding for improving resistance to stripe rust.

Photo-on-Demand Base-Catalyzed Phosgenation Reactions with Chloroform: Synthesis of Arylcarbonate and Halocarbonate Esters.

Carbonate esters are utilized as solvents and reagents for C1 building blocks in organic synthesis. This study reports a novel photo-on-demand in situ synthesis of carbonate esters with CHCl3 solutions containing a mixture of an aromatic or haloalkyl alcohol having relatively high acidity, and an organic base. We found that the acid-base interaction of the alcohol and base in the CHCl3 solution plays a key role in enabling the photochemical reaction. This reaction allows practical syntheses of diphenyl carbonate derivatives, haloalkyl carbonates, and polycarbonates, which are important chemicals and materials in industry.

Photo-on-Demand Synthesis of Vilsmeier Reagents with Chloroform and Their Applications to One-Pot Organic Syntheses.

The Vilsmeier reagent (VR), first reported a century ago, is a versatile reagent in a variety of organic reactions. It is used extensively in formylation reactions. However, the synthesis of VR generally requires highly toxic and corrosive reagents such as POCl3, SOCl2, or COCl2. In this study, we found that VR is readily obtained from a CHCl3 solution containing N,N-dimethylformamide or N,N-dimethylacetamide upon photo-irradiation under O2 bubbling. The corresponding Vilsmeier reagents were obtained in high yields with the generation of gaseous HCl and CO2 as byproducts to allow their isolations as crystalline solid products amenable to analysis by X-ray crystallography. With the advantage of using CHCl3, which bifunctionally serves as a reactant and a solvent, this photo-on-demand VR synthesis is available for one-pot syntheses of aldehydes, acid chlorides, formates, ketones, esters, and amides.

Modulation of the Aß-Peptide-Aggregation Pathway by Active Compounds From Platycladus orientalis Seed Extract in Alzheimer's Disease Models.

Alzheimer's disease (AD) is a neurodegenerative disease characterized by neuronal loss, cognitive impairment, and aphasia. Aggregation of ß-amyloid (Aß) peptide in the brain is considered a key mechanism in the development of AD. In the past 20 years, many compounds have been developed to inhibit Aß aggregation and accelerate its degradation. Platycladus orientalis seed is a traditional Chinese medicine used to enhance intelligence and slow aging. We previously found that Platycladus orientalis seed extract (EPOS) inhibited Aß-peptide aggregation in the hippocampus and reduced cognitive deficits in 5×FAD mice. However, the mechanisms of these effects have not been characterized. To characterize the protective mechanisms of EPOS, we used a transgenic Caenorhabditis elegans CL4176 model to perform Bioactivity-guided identification of active compounds. Four active compounds, comprising communic acid, isocupressic acid, imbricatolic acid, and pinusolide, were identified using 13C-and 1H-NMR spectroscopy. Furthermore, we showed that isocupressic acid inhibited Aß generation by modulating BACE1 activity via the GSK3ß/NF-κB pathway in HEK293-APPsw cells. These findings showed that EPOS reduced cognitive deficits in an AD model via modulation of the Aß peptide aggregation pathway.

Photo-on-Demand Synthesis of Chloroformates with a Chloroform Solution Containing an Alcohol and Its One-Pot Conversion to Carbonates and Carbamates.

Chloroformates are key reagents for synthesizing carbonates and carbamates. The present study reports a novel photo-on-demand in situ synthesis of chloroformates with a CHCl3 solution containing a primary alkyl alcohol. It further allowed the one-pot synthesis of carbonates and carbamates through subsequent addition of alcohols or amines, respectively.

Hydroxychalcone dyes that serve as color indicators for pH and fluoride ions.

A chalcone, which is composed of two aromatic rings bridged by an α,ß-unsaturated carbonyl group, exhibits a variety of biological activities. With an objective to develop a novel chalcone-based functional dye, we have synthesized a chalcone diol CLN1, bearing two OH groups at the 2-positions on both phenyl rings, as well as reference compounds CLN2-6, and found that it serves as color indicators for pH and fluoride ions. CLN1 showed a vivid color change from colorless to yellow (halochromism) in water at pH ≥ 10. Furthermore, it presented a selective color change from colorless to red upon the addition of TBAF in an organic solvent such as CH3CN. CLN1 provided a strong red-shifted absorption band in the visible region under alkaline conditions in water and upon the addition of TBAF in CH3CN. The absorption spectral study together with TD-DFT calculations and X-ray crystallographic analysis revealed that the characteristic π-resonant structures of CLN1 caused by the ionization or OH-F- interactions and the planar conformation due to its intramolecular hydrogen bonding may provide a strong charge transfer (CT) absorption in the visible region.

Presence and function of stress granules in atrial fibrillation.

AIMS: Stress granules (SGs) are transient cytoplasmic mRNA and protein complexes that form in eukaryotic cells under stress. SGs are related to multiple diseases, but there are no reports of the existence of SGs in atrial fibrillation (AF). METHODS AND RESULTS: Cell models of AF were established by field stimulation at 600 times per minute. HL-1 cells, cardiomyocytes and cardiac fibroblasts were transfected with G3BP1-cDNA plasmid by Lipofectamine 2000. The presence of SGs was detected by immunofluorescence analysis against GTPase-activating protein SH3 domain binding protein 1 (G3BP1) and poly(A)-binding protein 1 (PABP-1) and electron microscopy. Stable HL-1 cell lines transfected with lentivirus overexpressing G3BP1were constructed to further induce the formation of SGs in AF. Reactive oxygen species (ROS) and calcium overload in tachypaced HL-1 cells were studied by flow cytometry. The effects of G3BP1 overexpression on cardiac fibroblast proliferation and the protein expression level of collagen I/III and fibronectin-1 were also studied. Additionally, we detected protein synthesis in general and in single cells by puromycin incorporation in paced HL-1 cells. Here, we first showed that SGs are present in both tachypaced mouse cardiomyocytes and HL-1 atrial cells, although the presence is partial and at a low level. G3BP1 overexpression promoted SG formation, inhibited the rapid pacing-induced increase in ROS level, and attenuated calcium overload in HL-1 cells (all P<0.05). Furthermore, G3BP1 overexpression inhibited cardiac fibroblast proliferation (P<0.05) and decreased the protein expression level of collagen I and fibronectin-1 in cardiac fibroblasts stimulated by angiotensin II (all P<0.05). The bulk puromycin incorporation analyzed by Western blot did not show a global reduction in protein synthesis. However, puromycin incorporation in single cells detected by immunofluorescence analysis showed that protein synthesis in SG-containing cells significantly reduced (P<0.01). CONCLUSIONS: SGs are rapidly induced and present partially in AF, and G3BP1 overexpression promotes SG formation and confers cytoprotection against oxidative stress, calcium overload and atrial fibrosis in AF.

Voluntary Exercise Promotes Glymphatic Clearance of Amyloid Beta and Reduces the Activation of Astrocytes and Microglia in Aged Mice.

Age is characterized by chronic inflammation, leading to synaptic dysfunction and dementia because the clearance of protein waste is reduced. The clearance of proteins depends partly on the permeation of the blood-brain barrier (BBB) or on the exchange of water and soluble contents between the cerebrospinal fluid (CSF) and the interstitial fluid (ISF). A wealth of evidence indicates that physical exercise improves memory and cognition in neurodegenerative diseases during aging, such as Alzheimer's disease (AD), but the influence of physical training on glymphatic clearance, BBB permeability and neuroinflammation remains unclear. In this study, glymphatic clearance and BBB permeability were evaluated in aged mice using in vivo two-photon imaging. The mice performed voluntary wheel running exercise and their water-maze cognition was assessed; the expression of the astrocytic water channel aquaporin 4 (AQP4), astrocyte and microglial activation, and the accumulation of amyloid beta (Aß) were evaluated with immunofluorescence or an enzyme-linked immunosorbent assay (ELISA); synaptic function was investigated with Thy1-green fluorescent protein (GFP) transgenic mice and immunofluorescent staining. Voluntary wheel running significantly improved water-maze cognition in the aged mice, accelerated the efficiency of glymphatic clearance, but which did not affect BBB permeability. The numbers of activated astrocytes and microglia decreased, AQP4 expression increased, and the distribution of astrocytic AQP4 was rearranged. Aß accumulation decreased, whereas dendrites, dendritic spines and postsynaptic density protein (PSD95) increased. Our study suggests that voluntary wheel running accelerated glymphatic clearance but not BBB permeation, improved astrocytic AQP4 expression and polarization, attenuated the accumulation of amyloid plaques and neuroinflammation, and ultimately protected mice against synaptic dysfunction and a decline in spatial cognition. These data suggest possible mechanisms for exercise-induced neuroprotection in the aging brain.

Transgenic over-expression of slit2 enhances disruption of blood-brain barrier and increases cell death after traumatic brain injury in mice.

Traumatic brain injury (TBI) is the leading cause of mortality and disability among male adolescents and young adults; and mild traumatic brain injury is the most common type of traumatic brain injury. The disruption of blood-brain barrier (BBB) plays an important role in brain trauma. Previously, we have found that slit2, a member of slit protein family, increases permeability of BBB. In the present study, we examined the role of slit2 in the pathogenesis of mild TBI in a mouse model of micro TBI. Rhodamine BandPI (PropidiumIodide) staining were used to detect the permeability of BBB and cell death, respectively. The leakage of Rhodamine B and cell death were significantly increased in Slit2-Tg mice than in C57 control mice after micro TBI. The present results suggest that over expression of slit2 plays a detrimental role in the pathophysiology of mild TBI.

Deferoxamine inhibits microglial activation, attenuates blood-brain barrier disruption, rescues dendritic damage, and improves spatial memory in a mouse model of microhemorrhages.

Cerebral microbleeds are strongly linked to cognitive dysfunction in the elderly. Iron accumulation plays an important role in the pathogenesis of intracranial hemorrhage. Deferoxamine (DFX), a metal chelator, removes iron overload and protects against brain damage in intracranial hemorrhage. In this study, the protective effects of DFX against microhemorrhage were examined in mice. C57BL6 and Thy-1 green fluorescent protein transgenic mice were subjected to perforating artery microhemorrhages on the right posterior parietal cortex using two-photon laser irradiation. DFX (100 mg/kg) was administered 6 h after microhemorrhage induction, followed by every 12 h for three consecutive days. The water maze task was conducted 7 days after induction of microhemorrhages, followed by measurement of blood-brain barrier permeability, iron deposition, microglial activation, and dendritic damage. Laser-induced multiple microbleeds in the right parietal cortex clearly led to spatial memory disruption, iron deposits, microglial activation, and dendritic damage, which were significantly attenuated by DFX, supporting the targeting of iron overload as a therapeutic option and the significant potential of DFX in microhemorrhage treatment. Irons accumulation after intracranial hemorrhage induced a serious secondary damage to the brain. We proposed that irons accumulation after parietal microhemorrhages impaired spatial cognition. After parietal multiple microhemorrhages, increased irons and ferritin contents induced blood-brain barrier disruption, microglial activation, and further induced dendrites loss, eventually impaired the water maze, deferoxamine treatment protected from these damages.

Rhizoma Anemarrhenae extract ameliorates hyperglycemia and insulin resistance via activation of AMP-activated protein kinase in diabetic rodents.

ETHNOPHARMACOLOGICAL RELEVANCE: Rhizoma Anemarrhenae has been used in Asian countries for thousands of years to treat diabetes. Insulin resistance (IR) is the primary cause responsible for type 2 diabetes. The aim of this study was to to assess the hypoglycemic and insulin sensitizing properties of Rhizoma Anemarrhenae extract (TFA) in animal models of insulin resistance and/or diabetes and to delineate modes of action. MATERIALS AND METHODS: In-vivo studies were performed on STZ-induced diabetic mice and KK-Ay mice, the former of which were given the extract alone or in combination with insulin for 7 days, and the latter of which were given the extract for 8 consecutive weeks. Fasting blood glucose and serum insulin levels were measured. Pancreatic tissue sections were examined using transmission electron micrographs. Further, hyperinsulinemic-euglycemic clamping study was conducted in BCG vaccine-induced insulin resistance rats, and glucose infusion rate was examined. Mechanisms of action were investigated in 3T3-L1 and Hela cells using Western blot analysis. RESULTS: Our study showed that TFA enhanced the glucose-lowering effects of exogenous insulin administration in STZ-induced diabetic mice. Therapeutic administration of TFA significantly reduced fasting blood glucose, and serum insulin levels, and markedly increased the size and the number of insulin-producing beta cells in KK-Ay mice. Further, hyperinsulinemic-euglycemic clamping study showed that glucose infusion rate was significantly improved in TFA-treated BCG vaccine-induced insulin resistance rats. Study of mechanism of action revealed that TFA increased phosphorylation of AMPK and its downstream target, acetyl-CoA carboxylase (ACC) in 3T3-L1 cells. It activates AMPK in a LKB1-independent manner, providing a unified explanation for the beneficial effects of TFA. CONCLUSIONS: This study that TFA mediates activation of AMPK and improves overall glucose and lipid metabolism in diabetic rodents, highlights the potential utility of TFA for the management of type 2 diabetes.