Dual roles of myeloid-derived suppressor cells in various diseases: a review.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of cells that originate from bone marrow stem cells. In pathological conditions, such as autoimmune disorders, allergies, infections, and cancer, normal myelopoiesis is altered to facilitate the formation of MDSCs. MDSCs were first shown to promote cancer initiation and progression by immunosuppression with the assistance of various chemokines and cytokines. Recently, various studies have demonstrated that MDSCs play two distinct roles depending on the physiological and pathological conditions. MDSCs have protective roles in autoimmune disorders (such as uveoretinitis, multiple sclerosis, rheumatoid arthritis, ankylosing spondylitis, type 1 diabetes, autoimmune hepatitis, inflammatory bowel disease, alopecia areata, and systemic lupus erythematosus), allergies, and organ transplantation. However, they play negative roles in infections and various cancers. Several immunosuppressive functions and mechanisms of MDSCs have been determined in different disease conditions. This review comprehensively discusses the associations between MDSCs and various pathological conditions and briefly describes therapeutic approaches.

Beyond the Diagnosis: A Deep Dive Into the End Stage Liver Disease Experience From the Patient Perspective.

INTRODUCTION: End-stage liver disease (ESLD) presents a multifaceted challenge that encompasses not only physical but also emotional, psychological, and social dimensions. This study aims to explore the experiences of ESLD patients within the United States healthcare system. METHODS: Utilizing a convenience sampling methodology, 15 ESLD patients from a tertiary care hospital in the USA participated in semi-structured interviews between April 2023 and January 2024. Data analysis was conducted using MAXQDA 2023, employing a phenomenological approach to identify common themes. RESULTS: The study identified six primary themes: the significance of communication style in diagnosis delivery, the crucial role of family and social support, varied understanding and preferences for palliative care, diverse attitudes towards advanced care planning, preferences for coordinated healthcare experiences, and the emotional and psychological impact of ESLD. CONCLUSION: Our study underscores the complexity of ESLD patient care beyond medical treatment, highlighting the importance of clear communication, empathetic care, and the integration of family and palliative care services.

A convenient spectrophotometric test for screening skin-sensitizing chemicals using reactivity with glutathione in chemico.

To initiate skin sensitization, haptens react with endogenous proteins. During this process, skin sensitizers react with small endogenous molecules containing thiol or amino groups. In this study, a simple spectrophotometric method to identify skin sensitizers in chemico was developed using the reactivity of glutathione (GSH) with test chemicals in a 96-well plate. To quantitate the remaining GSH following the reaction with a skin sensitizer, 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) was employed. The optimized experimental conditions included the pH- and time-dependent stability of GSH, stability of derivatized products of GSH with optimal concentration and incubation time of DTNB, incubation time of GSH with the test chemicals, and molar ratios of GSH to the test chemicals. With the optimized conditions with both acetonitrile and DMSO as vehicles and incubation of GSH with test chemicals in 1:10 and 1:15 ratios for 24 h at 4 °C, 23 skin sensitizers and 23 non-sensitizers, based on the local lymph node assay, were tested to determine the predictive capacity of individual conditions. The best result showed a predictive capacity of 95.2% sensitivity, 91.3% specificity, and 93.2% accuracy, with 93.2% consistency in three trials, when 5.8% depletion was used as a cut-off value in 1:10 of GSH:test chemicals in DMSO. It would be an economic and useful screening tool for determining the skin sensitization potential of small molecules, because the present method employs simple endogenous GSH as an electron donor for sensitizers with a spectrophotometric detection system in 96-well plates, and because the method requires neither experimental animals nor cell cultures. Supplementary Information: The online version contains supplementary material available at 10.1007/s43188-023-00218-9.

Targeting a xenobiotic transporter to ameliorate vincristine-induced sensory neuropathy.

Vincristine is a widely used chemotherapeutic drug for the treatment of multiple malignant diseases that causes a dose-limiting peripheral neurotoxicity. There is no clinically effective preventative treatment for vincristine-induced sensory peripheral neurotoxicity (VIPN), and mechanistic details of this side effect remain poorly understood. We hypothesized that VIPN is dependent on transporter-mediated vincristine accumulation in dorsal root ganglion neurons. Using a xenobiotic transporter screen, we identified OATP1B3 as a neuronal transporter regulating the uptake of vincristine. In addition, genetic or pharmacological inhibition of the murine orthologue transporter OATP1B2 protected mice from various hallmarks of VIPN - including mechanical allodynia, thermal hyperalgesia, and changes in digital maximal action potential amplitudes and neuronal morphology - without negatively affecting plasma levels or antitumor effects of vincristine. Finally, we identified α-tocopherol from an untargeted metabolomics analysis as a circulating endogenous biomarker of neuronal OATP1B2 function, and it could serve as a companion diagnostic to guide dose selection of OATP1B-type transport modulators given in combination with vincristine to prevent VIPN. Collectively, our findings shed light on the fundamental basis of VIPN and provide a rationale for the clinical development of transporter inhibitors to prevent this debilitating side effect.

Targeting OCT2 with Duloxetine to Prevent Oxaliplatin-Induced Peripheral Neurotoxicity.

Oxaliplatin-induced peripheral neurotoxicity (OIPN) is a debilitating side effect that afflicts ~90% of patients that is initiated by OCT2-dependent uptake of oxaliplatin in DRG neurons. The antidepressant drug duloxetine has been used to treat OIPN, although its usefulness in preventing this side effect remains unclear. We hypothesized that duloxetine has OCT2-inhibitory properties and can be used as an adjunct to oxaliplatin-based regimens to prevent OIPN. Transport studies were performed in cells stably transfected with mouse or human OCT2 and in isolated mouse DRG neurons ex vivo. Wild-type and OCT2-deficient mice were used to assess effects of duloxetine on hallmarks of OIPN, endogenous OCT2 biomarkers, and the pharmacokinetics of oxaliplatin, and the translational feasibility of a duloxetine-oxaliplatin combination was evaluated in various models of colorectal cancer. We found that duloxetine potently inhibited the OCT2-mediated transport of several xenobiotic substrates, including oxaliplatin, in a reversible, concentration-dependent manner, and independent of species and cell context. Furthermore, duloxetine restricted access of these substrates to DRG neurons ex vivo and prevented OIPN in wild-type mice to a degree similar to the complete protection observed in OCT2-deficient mice, without affecting the plasma levels of oxaliplatin. Importantly, the uptake and cytotoxicity of oxaliplatin in tumor cell lines in vitro and in vivo were not negatively influenced by duloxetine. The observed OCT2-targeting properties of duloxetine, combined with the potential for clinical translation, provide support for its further exploration as a therapeutic candidate for studies aimed at preventing OIPN in cancer patients requiring treatment with oxaliplatin. Significance: We found that duloxetine has potent OCT2-inhibitory properties and can diminish excessive accumulation of oxaliplatin into DRG neurons. In addition, pre-treatment of mice with duloxetine prevented OIPN without significantly altering the plasma pharmacokinetics and antitumor properties of oxaliplatin. These results suggest that intentional inhibition of OCT2-mediated transport by duloxetine can be employed as a prevention strategy to ameliorate OIPN without compromising the effectiveness of oxaliplatin-based treatment.

Engineering of hybrid spheroids of mesenchymal stem cells and drug depots for immunomodulating effect in islet xenotransplantation.

Immunomodulation is an essential consideration for cell replacement procedures. Unfortunately, lifelong exposure to nonspecific systemic immunosuppression results in immunodeficiency and has toxic effects on nonimmune cells. Here, we engineered hybrid spheroids of mesenchymal stem cells (MSCs) with rapamycin-releasing poly(lactic-co-glycolic acid) microparticles (RAP-MPs) to prevent immune rejection of islet xenografts in diabetic C57BL/6 mice. Hybrid spheroids were rapidly formed by incubating cell-particle mixture in methylcellulose solution while maintaining high cell viability. RAP-MPs were uniformly distributed in hybrid spheroids and sustainably released RAP for ~3 weeks. Locoregional transplantation of hybrid spheroids containing low doses of RAP-MPs (200- to 4000-ng RAP per recipient) significantly prolonged islet survival times and promoted the generation of regional regulatory T cells. Enhanced programmed death-ligand 1 expression by MSCs was found to be responsible for the immunomodulatory performance of hybrid spheroids. Our results suggest that these hybrid spheroids offer a promising platform for the efficient use of MSCs in the transplantation field.

MATE1 Deficiency Exacerbates Dofetilide-Induced Proarrhythmia.

Dofetilide is a rapid delayed rectifier potassium current inhibitor widely used to prevent the recurrence of atrial fibrillation and flutter. The clinical use of this drug is associated with increases in QTc interval, which predispose patients to ventricular cardiac arrhythmias. The mechanisms involved in the disposition of dofetilide, including its movement in and out of cardiomyocytes, remain unknown. Using a xenobiotic transporter screen, we identified MATE1 (SLC47A1) as a transporter of dofetilide and found that genetic knockout or pharmacological inhibition of MATE1 in mice was associated with enhanced retention of dofetilide in cardiomyocytes and increased QTc prolongation. The urinary excretion of dofetilide was also dependent on the MATE1 genotype, and we found that this transport mechanism provides a mechanistic basis for previously recorded drug-drug interactions of dofetilide with various contraindicated drugs, including bictegravir, cimetidine, ketoconazole, and verapamil. The translational significance of these observations was examined with a physiologically-based pharmacokinetic model that adequately predicted the drug-drug interaction liabilities in humans. These findings support the thesis that MATE1 serves a conserved cardioprotective role by restricting excessive cellular accumulation and warrant caution against the concurrent administration of potent MATE1 inhibitors and cardiotoxic substrates with a narrow therapeutic window.

Outcomes, Trends, and Predictors of Gastrointestinal Bleeding in Patients Undergoing Transcatheter Aortic Valve Implantation (from the National Inpatient Sample).

Major bleeding has been identified as one of the most common complications after transcatheter aortic valve implantation (TAVI) with some suffering gastrointestinal bleeding (GIB). This study aimed at assessing the incidence and predictors of GIB after TAVI in the United States. We performed a retrospective analysis of data from the National Inpatient Sample database from 2011 to 2018. A total of 216,023 hospitalizations for TAVI were included. Of the included patients, 2,188 (1%) patients had GIB, whereas 213,835 (99%) patients did not have GIB. The presence of arteriovenous malformation was associated with the highest odds of having a gastrointestinal bleed (odds ratio (OR) 24.8, 95% confidence interval (CI) 17.13 to 35.92). Peptic ulcer disease was associated with an eightfold increased risk of bleeding (OR 8.74, 95% CI, 6.69 to 11.43) followed closely by colorectal cancer (OR 7.89, 95% CI, 5.33 to 11.70). Other comorbidities that were associated with higher propensity-matched rates of GIB were chronic kidney disease (OR 1.27,95% CI, 1.14 to 1.41), congestive heart failure (OR 1.18, 95% CI,1.06 to 1.32), liver disease (OR1.83, 95% CI,1.53 to 2.19), end-stage renal disease (OR 2.08,95% CI, 1.75 to 2.47), atrial fibrillation (OR1.63,95% CI,  1.49 to 1.78), and lung cancer (OR 2.80, 95% CI,1.77 to 4.41). Patients with GIB had higher propensity-matched rates of mortality than those without GIB, (12.1% vs 3.2%, p <0.01). Patients with GIB had a higher median cost of stay ($68,779 vs $46,995, p <0.01) and a longer length of hospital stay (11 vs 3 days, p <0.01). In conclusion, health care use and mortality are higher in hospitalizations of TAVI with a GIB. Baseline comorbidities like peptic ulcer disease, chronic kidney disease, liver disease, atrial fibrillation and, colorectal cancer are significant predictors of this adverse event.

Paraneoplastic Raynaud's Phenomenon as an Initial Manifestation of Lung Cancer?

Paraneoplastic Raynaud's phenomenon has often been reported in leukaemia, lymphoma and sarcoma. Nevertheless, an association with lung cancer is not frequently reported in the literature even though lung cancer is a common malignancy. We present a case of paraneoplastic Raynaud's phenomenon as the presenting feature of underlying lung malignancy. LEARNING POINTS: Raynaud's phenomenon can be a presenting feature of lung cancer.Evaluation for an underlying malignancy is important if the work-up is negative for autoimmune and vascular aetiology.

A Rare Case of Serrated Polyposis Syndrome with the MSH6 and SMARCA4 variants.

Serrated polyposis syndrome (SPS) is the most common form of polyposis syndrome and has been shown to increase the risk of colorectal cancer (CRC). The genetic pathway of CRC in SPS is different from the classic adenomatous polyposis coli (APC) pathway, which accounts for 70-80% of cases of CRC. Most commonly, SPS mutations include BRAF and KRAS, with activation of the RAS-RAF-MAP kinase pathway involved in the pathogenesis of serrated lesions. We present a rare case of SPS in a 32-year-old woman with MSH6 and SMARCA4 variants, which have not previously been reported in the literature. LEARNING POINTS: Patients with serrated polyposis syndrome should receive frequent colon cancer screening.Patients and their relatives should undergo surveillance.

A convenient fluorometric test method for skin sensitization using glutathione in chemico.

A convenient fluorometrical test method to identify skin sensitizers in chemico was developed using reactivity with glutathione (GSH), a low molecular weight endogenous substance. Following incubation of test chemicals with GSH, the remaining GSH was quantitated fluorometrically by using monobromobimane (mBBr), a thiol-detecting agent, for determining % depletion of this endogenous substance by test chemicals. The experimental conditions optimized were: (1) reactivity of thiol compounds including GSH with mBBr, (2) effects of vehicles on reactivity, (3) molar ratios of GSH to test chemicals, and (4) reactivity of endogenous substance with test substances under different incubation times. When an optimized condition with DMSO as a vehicle for test chemicals and in 1:60 ratio for 24 hr at 4°C was applied to classify 48 well-known skin sensitizers and non-sensitizers, the predictive capacity was as follows: 88.2% sensitivity, 78.6% specificity, and 85.4% accuracy with 95.8% consistency of three trials when 10.3% depletion of GSH was used as a cutoff value. Because the present method employed relatively simple GSH as an acceptor for sensitizers and/or a relatively convenient fluorometric detection system in 96-well plates for a high throughput test, it would be a useful test tool for screening skin sensitization potential of test chemicals.

Effects of Intestinal Microbiota on Pharmacokinetics of Crocin and Crocetin in Male Sprague-Dawley Rats.

In addition to the hepatic metabolism, the role of intestinal microbiota in drug metabolism has been considered important in the biotransformation of xenobiotics. Crocin and its aglycone, crocetin, isolated from many plants, including the dried stigma of Crocus sativus and the fruit of Gardenia jasminoides, have been used in treatment of inflammation, cancer, and metabolic disorders. In this study, the effect of intestinal microbiota on the pharmacokinetics of crocin was studied following single oral treatment with 600 mg/kg crocin to male rats pre-treated with a mixture of antibiotics, such as cefadroxil, oxytetracycline, and erythromycin, for three consecutive days. Following crocin treatment, blood, urine, and feces were collected at various time points for evaluating pharmacokinetic characteristics of crocin and crocetin by using LC-MS. Results showed that intestinal absorption of crocin was relatively marginal when compared with that of crocetin, and that crocin metabolism to crocetin by intestinal microbiota would be a critical step for absorption. The present results clearly suggested that the in vivo pharmacological effects of crocin might be considered as the effects by its aglycone, crocetin, mainly, and that the metabolism of glycosidic natural products by intestinal microbiota should be considered to understand their pharmacodynamic actions.

Role of Intestinal Microbiota in Metabolism of Voglibose In Vitro and In Vivo.

BACKGROUND: Voglibose, an α-glucosidase inhibitor, inhibits breakdown of complex carbohydrates into simple sugar units in intestine. Studies showed that voglibose metabolism in the liver might be negligible due to its poor intestinal absorption. Numerous microorganisms live in intestine and have several roles in metabolism and detoxification of various xenobiotics. Due to the limited information, the possible metabolism of voglibose by intestinal microbiota was investigated in vitro and in vivo. METHODS: For the in vitro study, different concentrations of voglibose were incubated with intestinal contents, prepared from both vehicle- and antibiotics-treated mice, to determine the decreased amount of voglibose over time by using liquid chromatography-mass spectrometry. Similarly, in vivo pharmacodynamic effect of voglibose was determined following the administration of voglibose and starch in vehicle- and antibiotic-pretreated non-diabetic and diabetic mice, by measuring the modulatory effects of voglibose on blood glucose levels. RESULTS: The in vitro results indicated that the remaining voglibose could be significantly decreased when incubated with the intestinal contents from normal mice compared to those from antibiotic-treated mice, which had less enzyme activities. The in vivo results showed that the antibiotic pretreatment resulted in reduced metabolism of voglibose. This significantly lowered blood glucose levels in antibiotic-pretreated mice compared to the control animals. CONCLUSION: The present results indicate that voglibose would be metabolized by the intestinal microbiota, and that this metabolism might be pharmacodynamically critical in lowering blood glucose levels in mice.

Alternative Methods for Testing Botulinum Toxin: Current Status and Future Perspectives.

Botulinum toxins are neurotoxic modular proteins composed of a heavy chain and a light chain connected by a disulfide bond and are produced by Clostridium botulinum. Although lethally toxic, botulinum toxin in low doses is clinically effective in numerous medical conditions, including muscle spasticity, strabismus, hyperactive urinary bladder, excessive sweating, and migraine. Globally, several companies are now producing products containing botulinum toxin for medical and cosmetic purposes, including the reduction of facial wrinkles. To test the efficacy and toxicity of botulinum toxin, animal tests have been solely and widely used, resulting in the inevitable sacrifice of hundreds of animals. Hence, alternative methods are urgently required to replace animals in botulinum toxin testing. Here, the various alternative methods developed to test the toxicity and efficacy of botulinum toxins have been briefly reviewed and future perspectives have been detailed.

Inflammation-triggered local drug release ameliorates colitis by inhibiting dendritic cell migration and Th1/Th17 differentiation.

Enteric-coated formulations using Eudragit® polymers have been extensively used for delivering drugs to the lower gastrointestinal tract. However, these drug-delivery systems cannot accurately deliver the therapeutic cargoes to colon because of early degradation of the polymers at alkaline pH of the small intestine. Here, we describe a precise method of delivering drugs to inflammation sites in colon using an oral drug delivery system. Tacrolimus (FK506)-loaded microspheres were prepared using a thioketal-based polymer that releases drug in response to reactive oxygen species (ROS), which are abundantly produced at the sites of inflammation in acute colitis. Orally-administered FK506-loaded thioketal microspheres (FK506-TKM) led to a substantial accumulation of FK506 in inflamed colon and effectively alleviated dextran-sulfate sodium (DSS)-induced murine colitis. At the molecular level, FK506-TKM significantly inhibited infiltration of CD4+ and CD8+ T lymphocytes in colon and differentiation of CD4+ T cells into Th1 and Th17 cells in colon-draining mesenteric lymph nodes via restricting dendritic cell migration from colon. Our findings indicate orally-administered thioketal-based drug delivery system as a promising means of treating acute inflammatory bowel diseases.

Assessment of skin sensitizing potential of metals with ß-galactosidase-expressing E. coli culture system.

It has been a challenge to develop in vitro alternative test methods for accurate prediction of metallic products which may exert skin sensitization, as several test methods adopted by OECD were relatively ineffective in assessing the capacity for metallic compounds to exert sensitizing reactions, compared with organic test substances. Based upon these findings, a system that incorporates ß-galactosidase producing E. coli cultures was tested for its predictive capacity to well-known metallic sensitizers. In this system, E. coli cells were incubated with metal salts at various concentrations and ß-galactosidase suppression by each test metal was determined. Fourteen local lymph node assay (LLNA) categorized metal salts were examined. Although color interference from metal salts was minimal, a fluorometric detection system was also employed using 4-methylumbelliferyl galactopyranoside as a substrate for ß-galactosidase to avoid the color interference, concomitantly with the original UV-spectrometric method. Data demonstrated that two detection methods were comparable and complementary. In addition, most of the metallic sensitizers were correctly identified at 0.6 and 0.8 mM concentrations. Despite the lower specificity obtained in the current study and small number of substances tested, the developed method appears to be a relatively simple and effective in vitro method for detecting metallic sensitizers. When 61 chemicals tested in the ß-galactosidase producing E. coli cultures including the present study were collectively analyzed, the prediction capacity was as high as other OECD-adopted tests: 95.6% of sensitivity, 66.7% of specificity, and 88.5% of accuracy. It is important to emphasize that animals or mammalian cell cultures were not required in the current method, which are in accordance with the EU guidelines on restricted or banned animal testing.

Engineering "cell-particle hybrids" of pancreatic islets and bioadhesive FK506-loaded polymeric microspheres for local immunomodulation in xenogeneic islet transplantation.

Host immune response remains an obstacle in cell-replacement therapy for treating type I diabetes. Long-term systemic immunosuppression results in suboptimal efficacy and adverse reactions. Thus, "cell-particle hybrids" of pancreatic islets and tissue-adhesive, polydopamine-coated, FK506-loaded biodegradable microspheres (PD-FK506-MS) were developed to locally modulate the immune response at the transplantation site. Coating of FK506-MS with PD enabled the rapid formation of stable cell-particle hybrids without significant changes in islet viability and functionality. Extremely low quantities of FK506 (approximately 600 ng per recipient) sustainably released from cell-particle hybrids effectively prolonged survival of xenogeneic islet graft. Interestingly, FK506 exhibited extended bioavailability in the grafts but was undetectable in systemic circulation and other tissues. Moreover, mRNA expression of inflammatory cytokines was significantly inhibited in the PD-FK506-MS-containing grafts but not in lymphoid organs. This study presents a promising platform that facilitates the translation of local immunomodulation towards an effective strategy with improved safety profiles for treating type I diabetes.

Discovery and Biological Evaluations of Halogenated 2,4-Diphenyl Indeno[1,2-b]pyridinol Derivatives as Potent Topoisomerase IIα-Targeted Chemotherapeutic Agents for Breast Cancer.

With the aim of developing new effective topoisomerase IIα-targeted anticancer agents, we synthesized a series of hydroxy- and halogenated 2,4-diphenyl indeno[1,2-b]pyridinols using a microwave-assisted single step synthetic method and investigated structure-activity relationships. The majority of compounds with chlorophenyl group at 2-position and phenol group at the 4-position of indeno[1,2-b]pyridinols exhibited potent antiproliferative activity and topoisomerase IIα-selective inhibition. Of the 172 compounds tested, 89 showed highly potent and selective topoisomerase IIα inhibition and antiproliferative activity in the nanomolar range against human T47D breast (2.6 nM) cancer cell lines. In addition, mechanistic studies revealed compound 89 is a nonintercalative topoisomerase II poison, and in vitro studies showed it had promising cytotoxic effects in diverse breast cancer cell lines and was particularly effective at inducing apoptosis in T47D cells. Furthermore, in vivo administration of compound 89 had significant antitumor effects in orthotopic mouse model of breast cancer.

Activation of Mevalonate Pathway via LKB1 Is Essential for Stability of Treg Cells.

The function of regulatory T (Treg) cells depends on lipid oxidation. However, the molecular mechanism by which Treg cells maintain lipid metabolism after activation remains elusive. Liver kinase B1 (LKB1) acts as a coordinator by linking cellular metabolism to substrate AMP-activated protein kinase (AMPK). We show that deletion of LKB1 in Treg cells exhibited reduced suppressive activity and developed fatal autoimmune inflammation. Mechanistically, LKB1 induced activation of the mevalonate pathway by upregulating mevalonate genes, which was essential for Treg cell functional competency and stability by inducing Treg cell proliferation and suppressing interferon-gamma and interleukin-17A expression independently of AMPK. Furthermore, LKB1 was found to regulate intracellular cholesterol homeostasis and to promote the mevalonate pathway. In agreement, mevalonate and its metabolite geranylgeranyl pyrophosphate inhibited conversion of Treg cells and enhanced survival of LKB1-deficient Treg mice. Thus, LKB1 is a key regulator of lipid metabolism in Treg cells, involved in optimal programming of suppressive activity, immune homeostasis, and tolerance.

Identification of DNA and glutathione adducts in male Sprague-Dawley rats exposed to 1-bromopropane.

Occupational exposure of workers to 1-bromopropane (1-BP) has raised concerns in industry for many years. Despite the known toxicity of this chemical, molecular events attributed to exposure to 1-BP have not been extensively studied. The aim of the present study was to examine the effects of 1-BP exposure on adduct formation with DNA and glutathione (GSH) in male Sprague-Dawley rats in an attempt to determine the early stages of toxicity. Following 6 h after either single or daily exposure to 1-BP for 3 days, N7-propyl guanine and S-propyl GSH were quantified in several organs by using liquid chromatography-mass spectrometry (LC-MS/MS). The results showed that N7-propyl guanine was maximally formed in liver followed by spleen, testes, and lung in both dose- and time-dependent manners. However, DNA adduct was not detected in cardiac tissue. In the case of S-propyl GSH, this compound was formed in the following order in various organs: liver > testes > spleen > kidney > lung > heart. In a subsequent in vitro study, formation of N7-propyl guanine initiated by 1-BP in calf thymus DNA was not markedly affected by addition of liver homogenates, which indicated that this chemical may be acting as a direct alkylating agent. In contrast, an in vitro study with free GSH demonstrated that 1-BP reduced GSH and elevated production of S-propyl GSH, and that the production of this adduct was significantly higher in the presence of active liver homogenates. Data indicated that formation of GSH adducts initiated by 1-BP might be associated with an enzyme-driven process. Although further characterization is necessary, it would appear that N7-propyl guanine and S-propyl GSH might serve as useful markers in cases of exposure assessment of 1-BP.